WO2022015115A1 - Therapeutic use of combination containing triple agonistic long-acting conjugate or triple agonist - Google Patents

Therapeutic use of combination containing triple agonistic long-acting conjugate or triple agonist Download PDF

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WO2022015115A1
WO2022015115A1 PCT/KR2021/009210 KR2021009210W WO2022015115A1 WO 2022015115 A1 WO2022015115 A1 WO 2022015115A1 KR 2021009210 W KR2021009210 W KR 2021009210W WO 2022015115 A1 WO2022015115 A1 WO 2022015115A1
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Prior art keywords
cysteine
glutamine
triple
pharmaceutical composition
lysine
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PCT/KR2021/009210
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French (fr)
Korean (ko)
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김정국
이종석
이상현
오의림
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한미약품 주식회사
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Priority to BR112023000585A priority Critical patent/BR112023000585A2/en
Priority to MX2023000699A priority patent/MX2023000699A/en
Priority to IL299841A priority patent/IL299841A/en
Priority to CN202180062143.3A priority patent/CN116209475A/en
Priority to US18/016,145 priority patent/US20230310630A1/en
Priority to CA3186199A priority patent/CA3186199A1/en
Priority to AU2021310793A priority patent/AU2021310793A1/en
Priority to EP21841859.8A priority patent/EP4183419A1/en
Priority to JP2023503210A priority patent/JP2023535382A/en
Publication of WO2022015115A1 publication Critical patent/WO2022015115A1/en

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    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
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    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
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    • A61K47/6801—Drug-antibody or immunoglobulin conjugates defined by the pharmacologically or therapeutically active agent
    • A61K47/6803—Drugs conjugated to an antibody or immunoglobulin, e.g. cisplatin-antibody conjugates
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    • A61P1/16—Drugs for disorders of the alimentary tract or the digestive system for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
    • A—HUMAN NECESSITIES
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    • A61K31/215—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids
    • A61K31/22—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids of acyclic acids, e.g. pravastatin
    • A61K31/222—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids of acyclic acids, e.g. pravastatin with compounds having aromatic groups, e.g. dipivefrine, ibopamine
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    • A61K31/34—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having five-membered rings with one oxygen as the only ring hetero atom, e.g. isosorbide
    • A61K31/343—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having five-membered rings with one oxygen as the only ring hetero atom, e.g. isosorbide condensed with a carbocyclic ring, e.g. coumaran, bufuralol, befunolol, clobenfurol, amiodarone
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    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4427—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems
    • A61K31/4436—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems containing a heterocyclic ring having sulfur as a ring hetero atom
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    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
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    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/56—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
    • A61K31/575—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids substituted in position 17 beta by a chain of three or more carbon atoms, e.g. cholane, cholestane, ergosterol, sitosterol
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    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A—HUMAN NECESSITIES
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    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/177—Receptors; Cell surface antigens; Cell surface determinants
    • A61K38/1796—Receptors; Cell surface antigens; Cell surface determinants for hormones
    • A—HUMAN NECESSITIES
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    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/26—Glucagons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/68—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/575—Hormones
    • C07K14/605—Glucagons
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • C07K14/72—Receptors; Cell surface antigens; Cell surface determinants for hormones
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    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2300/00—Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/30—Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto

Definitions

  • the present invention provides a triple agonistic long-acting conjugate or a triple agonist; and an FXR agonist (farnesoid X receptor agonist), or to the therapeutic use of the combination further comprising an acetyla-CoA carboxylase inhibitor (ACC) in the combination.
  • FXR agonist farnesoid X receptor agonist
  • liver-related diseases include nonalcoholic fatty liver, hepatitis, liver fibrosis, cholestatic liver disease, cirrhosis, liver failure, liver cancer, and the like.
  • Inflammation of the liver may occur due to causes such as viruses, alcohol, drugs, immune abnormalities, metabolic diseases, and the like, and it is known that diseases such as liver fibrosis, cirrhosis, liver cancer and liver cancer occur as hepatitis progresses and becomes chronic.
  • hepatitis which is caused by inflammation of the liver, accounts for most of the liver diseases, and it is known that various liver diseases (liver fibrosis, cirrhosis, etc.) appear due to liver inflammation or liver inflammation as hepatitis progresses.
  • liver diseases liver fibrosis, cirrhosis, etc.
  • acute hepatitis and chronic hepatitis depending on the cause, can be divided into viral hepatitis, alcoholic hepatitis, drug hepatitis, and the like.
  • Cholestatic liver disease (cholestasis liver disease) is also assumed to be caused by an inflammatory disease.
  • nonalcoholic fatty liver disease is a type of disease that shows tissue findings similar to alcoholic hepatitis even though it is not related to alcohol intake.
  • NASH nonalcoholic fatty liver disease
  • Nonalcoholic fatty liver disease is on the rise as the obesity and diabetic populations increase.
  • TZD-type drugs are not applicable to patients with heart disease because of the disadvantages of large weight gain and slowing the flow of body fluids.
  • GLP-1 receptor agonists such as Victoza or Byetta
  • the drug was expected to have a therapeutic effect on nonalcoholic fatty liver disease and fibrosis caused by it in the early stage of development because it is accompanied by a weight loss effect.
  • the half-life in the body is extremely short, and since repeated administration is required every day or twice a day, there is a disadvantage in patient convenience. Such frequent administration causes great pain and discomfort to the patient.
  • GLP-1 Glucagon-like peptide-1
  • GIP Glucose-dependent insuliontropic polypeptide
  • GLP-1 has potential as a therapeutic agent for obesity, and GIP functions to promote insulin secretion from the pancreas and to lower blood sugar concentration dependent on blood sugar concentration, and increases the activity of GLP-1, anti-inflammatory effects, etc.
  • Glucagon is produced by the pancreas when blood sugar begins to drop due to a cause, such as medication or disease, or a hormone or enzyme deficiency. Glucagon signals the liver to break down glycogen to release glucose, and plays a role in raising blood sugar levels to normal levels.
  • glucagon exhibits anti-obesity effects by promoting lipolysis and energy expenditure by activating hormone-sensitive lipase in adipocytes and suppressing appetite in animals and humans, in addition to raising blood sugar. This has been reported
  • FXR agonists are FXR agonists, which are nuclear receptors activated by bile acids, also known as bile acid receptors (BARs). FXR is expressed in major sites of bile acid metabolism such as liver, intestine and kidney, and it has been reported that it acts in a tissue-specific manner to affect several metabolic pathways including bile acids as well as fibrosis. Accordingly, there is a need for an effective treatment method for liver disease mediated by FXR.
  • ACC inhibitors are known as substances that inhibit acetyla-CoA carboxylase, an enzyme important in the regulation of fatty acid production and metabolism.
  • One object of the present invention is to treat or prevent liver disease containing a pharmaceutically effective amount of a triple agonistic long-acting conjugate or a triple agonist and a pharmaceutically acceptable excipient.
  • Another object of the present invention is (i) a substance having activity on glucagon receptor, GLP-1 (Glucagon-like peptide-1) receptor, and GIP (glucose-dependent insuliontropic polypeptide) receptor, or a long-acting conjugate thereof, and (ii) a farnesoid X receptor agonist (FXR agonist).
  • Another object of the present invention is (i) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor or a long-acting conjugate thereof, (ii) an FXR agonist, and (iii) an acetyl coenzyme A carboxylase To provide a combination comprising an inhibitor (acetyla-CoA carboxylase inhibitor, ACC inhibitor).
  • Another object of the present invention is to provide a pharmaceutical composition for preventing or treating liver disease, comprising the combination.
  • Another object of the present invention comprises (i) a substance having activity on glucagon receptor, GLP-1 receptor, and GIP receptor or a long-acting conjugate thereof, and (ii) FXR agonist (farnesoid X receptor agonist), To provide a pharmaceutical kit for the prevention or treatment of liver disease.
  • Another object of the present invention is comprising (i) a substance having activity on glucagon receptor, GLP-1 receptor, and GIP receptor or a long-acting conjugate thereof, (ii) an FXR agonist, and (iii) an ACC inhibitor, To provide a pharmaceutical kit for the prevention or treatment of liver disease.
  • Another object of the present invention is to treat liver disease comprising administering and/or using the combination, a pharmaceutical composition for the treatment or prevention of liver disease, or the pharmaceutical kit to an individual in need thereof. To provide a method of prevention or treatment.
  • Another object of the present invention is to provide use of the combination, pharmaceutical composition, or pharmaceutical kit for the prophylaxis or treatment of liver disease and/or use for the preparation of a medicament for the prevention or treatment of non-alcoholic fatty liver disease. will be.
  • the triple acting long-acting conjugate or triple agent according to the present invention and concomitant administration of an FXR agonist, or a triple acting long-acting conjugate or a triple agent; FXR agonists; And the triple combination administration regimen of an ACC inhibitor has an improved effect compared to a single administration regimen, so it can be usefully used for the prevention or treatment of liver disease.
  • FIG. 1 is a diagram showing the change in NAS (NAFLD activity score) according to the complex administration of the long-acting conjugate of the triple activator (*p ⁇ 0.05, **p ⁇ 0.01, ***p ⁇ 0.001 vs. CH-HFD , vehicle control by ono-way ANOVA).
  • FIG. 2 is a diagram showing the change in the content of hydroxyprolin in liver tissue according to the complex administration of the long-acting conjugate of the triple activator (*p ⁇ 0.05, **p ⁇ 0.01, ***p ⁇ 0.001 vs. CH-HFD, vehicle control by ono-way ANOVA).
  • One aspect embodying the present invention is a pharmaceutical composition for the treatment or prevention of liver disease comprising a triple agonistic long-acting conjugate or a triple agonist, the pharmaceutical composition
  • the red composition is a composition characterized in that it is used in combination with a farnesoid X receptor agonist.
  • the pharmaceutical composition contains a pharmaceutically effective amount of a triple-acting long-acting conjugate or a triple agonist and a pharmaceutically acceptable excipient for the treatment or prevention of liver disease.
  • a composition a pharmaceutically effective amount of a triple-acting long-acting conjugate or a triple agonist and a pharmaceutically acceptable excipient for the treatment or prevention of liver disease.
  • the pharmaceutical composition is characterized in that it is used in combination with a farnesoid X receptor agonist,
  • the triple-acting long-acting conjugate is characterized in that the pharmaceutical composition is a substance represented by the following formula (1):
  • Lx is a linker containing an ethylene glycol repeating unit, and x is 0 or a natural number
  • Fc is an immunoglobulin Fc region
  • Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 1,
  • Xaa1 is histidine (His, H), 4-imidazoacetyl (CA) or tyrosine (Tyr, Y),
  • Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid or 2-aminoisobutyric acid (Aib),
  • Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q),
  • Xaa7 is threonine (Thr, T) or isoleucine (Ile, I),
  • Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C) or valine (Val, V);
  • Xaa12 is lysine (Lys, K), serine (Ser, S) or isoleucine (Ile, I);
  • Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A) or cysteine (Cys, C);
  • Xaa14 is leucine (Leu, L), methionine (Met, M) or tyrosine (Tyr, Y);
  • Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E) or leucine (Leu, L),
  • Xaa16 is glycine (Gly, G), glutamic acid (Glu, E) or serine (Ser, S);
  • Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C) or lysine (Lys, K);
  • Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R) or histidine (His, H);
  • Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C) or valine (Val, V);
  • Xaa20 is lysine (Lys, K), glutamine (Gln, Q) or arginine (Arg, R);
  • Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C) or aspartic acid (Asp, D);
  • Xaa23 is isoleucine (Ile, I) or valine (Val, V),
  • Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D) or glutamic acid (Glu, E),
  • Xaa27 is valine (Val, V), leucine (Leu, L) or lysine (Lys, K);
  • Xaa28 is cysteine (Cys, C), lysine (Lys, K), alanine (Ala, A), asparagine (Asn, N) or aspartic acid (Asp, D);
  • Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q), threonine (Thr, T), glutamic acid (Glu, E) or histidine (His, H);
  • Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K) or histidine (His, H), or is absent;
  • R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107) or m-SSGQPPPS-n (SEQ ID NO: 108), or is absent;
  • m is -Cys-, -Pro- or -Gly-Pro-;
  • n is absent or is -Cys-, -Gly-, -Ser- or -His-Gly-.
  • Xaa2 is glycine, alpha-methyl-glutamic acid or 2-aminoisobutyric acid,
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine or valine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, glutamine or cysteine,
  • Xaa14 is leucine, tyrosine or methionine
  • Xaa15 is cysteine, leucine, glutamic acid or aspartic acid,
  • Xaa17 is glutamine, arginine, isoleucine, cysteine, glutamic acid or lysine,
  • Xaa18 is alanine, glutamine, arginine or histidine;
  • Xaa19 is alanine, glutamine, valine or cysteine
  • Xaa20 is lysine, arginine or glutamine
  • Xaa21 is glutamic acid, glutamine, leucine, cysteine or aspartic acid,
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid or aspartic acid;
  • Xaa27 is characterized in that it is leucine or lysine.
  • composition according to any one of the preceding embodiments, wherein the Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 2:
  • Xaa1 is 4-imidazoacetyl, histidine or tyrosine;
  • Xaa2 is glycine, alpha-methyl-glutamic acid or 2-aminoisobutyric acid;
  • Xaa10 is tyrosine or cysteine
  • Xaa13 is alanine, glutamine, tyrosine or cysteine,
  • Xaa14 is leucine, methionine or tyrosine
  • Xaa15 is aspartic acid, glutamic acid or leucine
  • Xaa16 is glycine, glutamic acid or serine,
  • Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine or lysine,
  • Xaa18 is alanine, glutamine, arginine or histidine;
  • Xaa19 is alanine, glutamine, cysteine or valine
  • Xaa20 is lysine, glutamine or arginine
  • Xaa21 is cysteine, glutamic acid, glutamine, leucine or aspartic acid,
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, alanine, glutamine, asparagine or glutamic acid,
  • Xaa28 is lysine, cysteine, asparagine or aspartic acid
  • Xaa29 is glycine, glutamine, cysteine or histidine
  • Xaa30 is cysteine, glycine, lysine or histidine
  • Xaa31 is proline or cysteine
  • Xaa40 is cysteine or absent.
  • composition according to any one of the preceding embodiments, wherein the Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 3:
  • Xaa1 is histidine or tyrosine
  • Xaa2 is alpha-methyl-glutamic acid or 2-aminoisobutyric acid
  • Xaa13 is alanine, tyrosine or cysteine
  • Xaa17 is arginine, cysteine or lysine
  • Xaa18 is alanine or arginine
  • Xaa19 is alanine or cysteine
  • Xaa21 is glutamic acid or aspartic acid
  • Xaa24 is glutamine or asparagine
  • Xaa28 is cysteine or aspartic acid
  • Xaa29 is cysteine, histidine or glutamine
  • Xaa30 is cysteine or histidine
  • Xaa31 is proline or cysteine
  • Xaa40 is cysteine or absent.
  • Xaa16 is glutamic acid
  • Xaa20 is lysine
  • the glutamic acid and the lysine form a lactam ring.
  • composition according to any one of the preceding embodiments, wherein the Z or triple agent is amidated at its C-terminus.
  • composition according to any one of the preceding embodiments, wherein the formula weight of the ethylene glycol repeating unit moiety in Lx is in the range of 1 to 100 kDa.
  • composition according to any one of the preceding embodiments, wherein the Z or triple agent is a peptide comprising one amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102.
  • composition according to any one of the preceding embodiments, wherein the Z or triple agent is SEQ ID NO: 21, 22, 42, 43, 50, 64, 66, 67, 70, 71, 76, 77, 96, 97 and It is characterized in that it is a peptide comprising an amino acid sequence selected from the group consisting of 100.
  • composition according to any one of the preceding embodiments, wherein the Z or triple agent is an amino acid selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 66, 67, 77, 96, 97 and 100 It is characterized in that it is a peptide comprising a sequence.
  • composition according to any one of the preceding embodiments, wherein the Z or triple agent is a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 77 and 96 do it with
  • the farnesoid X receptor agonist is cafestol (Cafestol), chenodeoxycholic acid, obeticholic acid, fexaramine ( Fexaramine), GW 4064, PX104, 6E-CDCA (6-ethyl-chedeoxycholic acid), AKN-083, Tropifexor, Cilofexor, EDP-305, AGN-242266, AGN-242256, EP- 024297, RDX-023, BWL-200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, PX20606, It is characterized in that at least one selected from the group consisting of EYP-001, TERN-101, TC-100 and INT-2228.
  • composition according to any one of the preceding embodiments, wherein the pharmaceutical composition is also used in combination with an acetyl-Coenzyme A carboxylase inhibitor.
  • acetyl coenzyme A carboxylase inhibitor is CP-640186, (4-piperidinyl)-piperazine derivatives ((4-piperidinyl)-piperazine derivatives), 1, 4-substituted cyclohexane derivatives, spirochromanone derivatives, spirolactam derivatives, spirodiamine derivatives, spiropentacylamide derivatives , Pseudopeptide pyrrolidinedione derivatives, Macrocyclic polyketone derivatives, Thiophene pyrimidone derivatives including firsocostat, amino-oxazole It is characterized in that at least one selected from the group consisting of amino-oxazole derivatives, azobenzimidazole derivatives and PF-05221304.
  • liver disease is non-alcoholic fatty liver disease or cholestatic liver disease.
  • nonalcoholic fatty liver disease is at least selected from the group consisting of steatosis simplex, liver inflammation, nonalcoholic fatty liver, nonalcoholic steatohepatitis, cirrhosis, liver fibrosis, liver failure, and liver cancer. It is characterized as a single disease.
  • composition according to any one of the preceding embodiments, wherein the cholestatic liver disease is any one selected from the group consisting of primary biliary cirrhosis, primary sclerosing cholangitis, and combinations thereof. characterized in that
  • nonalcoholic fatty liver disease is at least one disease selected from the group consisting of liver inflammation, nonalcoholic steatohepatitis and liver fibrosis.
  • Another aspect embodying the present invention is (i) a substance having activity against glucagon receptors, GLP-1 (Glucagon-like peptide-1) receptors, and GIP (Glucose-dependent insuliontropic polypeptide) receptors or long-acting conjugates thereof , and (ii) a farnesoid X receptor agonist.
  • the combination further comprises (iii) an acetyla-CoA carboxylase inhibitor.
  • the substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor is a peptide comprising the amino acid sequence represented by the general formula 1 described above. do.
  • Xaa14 is leucine or methionine
  • Xaa15 is characterized in that it is cysteine, aspartic acid, or leucine.
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine, or valine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, or cysteine
  • Xaa14 is leucine or methionine
  • Xaa15 is cysteine or aspartic acid
  • Xaa17 is glutamine, arginine, isoleucine, cysteine, or lysine;
  • Xaa18 is alanine, arginine, or histidine
  • Xaa19 is alanine, glutamine, or cysteine
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid, cysteine, or aspartic acid
  • Xaa23 is valine
  • Xaa24 is alanine, glutamine, cysteine, asparagine, or aspartic acid
  • Xaa27 is characterized in that it is leucine or lysine.
  • Xaa13 is alanine, tyrosine, or cysteine
  • Xaa15 is aspartic acid or glutamic acid
  • Xaa17 is glutamine, arginine, cysteine, or lysine
  • Xaa18 is alanine, arginine, or histidine
  • Xaa21 is cysteine, glutamic acid, glutamine, or aspartic acid
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, glutamine, or asparagine
  • Xaa28 is cysteine, asparagine, or aspartic acid
  • Xaa29 is glutamine, cysteine, or histidine
  • Xaa30 is characterized in that it is cysteine, lysine, or histidine.
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa7 is threonine
  • Xaa10 is tyrosine or cysteine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, or cysteine
  • Xaa14 is leucine or methionine
  • Xaa15 is cysteine or aspartic acid
  • Xaa16 is glutamic acid
  • Xaa17 is arginine, isoleucine, cysteine, or lysine;
  • Xaa18 is alanine, arginine, or histidine
  • Xaa19 is alanine, glutamine, or cysteine
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid or aspartic acid
  • Xaa23 is valine
  • Xaa24 is glutamine, asparagine, or aspartic acid
  • Xaa27 is leucine
  • Xaa28 is characterized in that it is cysteine, alanine, asparagine, or aspartic acid.
  • Xaa1 is histidine or 4-imidazoacetyl
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa3 is glutamine
  • Xaa7 is threonine
  • Xaa10 is tyrosine
  • Xaa12 is isoleucine
  • Xaa13 is alanine or cysteine
  • Xaa14 is methionine
  • Xaa15 is aspartic acid
  • Xaa16 is glutamic acid
  • Xaa17 is isoleucine or lysine
  • Xaa18 is alanine or histidine
  • Xaa19 is glutamine or cysteine
  • Xaa20 is lysine
  • Xaa21 is aspartic acid
  • Xaa23 is valine
  • Xaa24 is asparagine
  • Xaa27 is leucine
  • Xaa28 is alanine or asparagine
  • Xaa29 is glutamine or threonine
  • Xaa30 is characterized in that it is cysteine or lysine, or is absent.
  • the substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor is a peptide comprising the amino acid sequence of Formula 3 as described above.
  • R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), CSSGQPPPS (SEQ ID NO: 109), GPSSGAPPPS (SEQ ID NO: 110), GPSSGAPPPSC (SEQ ID NO: 111), PSSGAPPPS (SEQ ID NO: 112), PSSGAPPPSG (SEQ ID NO: 113), PSSGAPPPSHG (SEQ ID NO: 113) 114), PSSGAPPPSS (SEQ ID NO: 115), PSSGQPPPS (SEQ ID NO: 116), or PSSGQPPPSC (SEQ ID NO: 117), or is characterized in that it is absent.
  • the substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor is a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102 characterized.
  • the FXR agonist is cafestol, Chenodeoxycholic acid, Obeticholic acid, Fexaramine, GW 4064, PX104 , 6E-CDCA (6-ethyl-chedeoxycholic acid), AKN-083 and Tropifexor, Cilofexor, EDP-305 AGN-242266, AGN-242256, EP-024297, RDX-023, BWL- 200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, PX20606, EYP-001, TERN-101, It is characterized in that it is selected from the group consisting of TC-100 and INT-2228.
  • the ACC inhibitor is CP-640186, (4-piperidinyl)-piperazine derivatives, 1,4-substituted cyclohexane derivatives (1,4-disubstituted cyclohexane derivatives), Spirochromanone derivatives, Spirolactam derivatives, Spirodiamine derivatives, Spiropentacylamide derivatives, Pseudopeptide pyrrolidine Ionic derivatives (Pseudopeptide pyrrolidinedione derivatives), Macrocyclic polyketone derivatives (Macrocyclic polyketone derivatives), Thiophene pyrimidone derivatives including firsocostat, Amino-oxazole derivatives ), azobenzimidazole derivatives, and PF-05221304.
  • the conjugate is a glucagon receptor, a GLP-1 receptor, and a biocompatible substance capable of increasing the in vivo half-life of a substance having activity on the GIP receptor bound, long-acting It is characterized in that it is in the form of a conjugate.
  • the substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor is a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102,
  • the conjugate thereof is characterized in that the peptide and the immunoglobulin Fc region are linked through a linker, which is a non-peptide polymer.
  • Another embodiment embodying the present invention is a pharmaceutical composition for preventing or treating liver disease, comprising the combination.
  • the liver disease is characterized in that non-alcoholic fatty liver disease (non-alcoholic fatty liver disease) or cholestatic liver disease.
  • nonalcoholic fatty liver disease is selected from the group consisting of simple steatosis, liver inflammation, nonalcoholic fatty liver, nonalcoholic steatohepatitis, cirrhosis, liver fibrosis, liver failure and liver cancer. do it with
  • composition according to any one of the preceding embodiments, wherein the cholestatic liver disease is characterized in that it is any one selected from the group consisting of primary biliary cirrhosis, primary sclerosing cholangitis, and combinations thereof.
  • glucagon receptor GLP-1 (Glucagon-like peptide-1) receptor
  • GIP glycose-dependent insuliontropic
  • Polypeptide is a pharmaceutical composition for preventing or treating liver disease, including a substance having activity on the receptor or a conjugate thereof.
  • the composition is characterized in that it is further used in combination with an ACC inhibitor (acetyla-CoA carboxylase inhibitor).
  • an ACC inhibitor acetyla-CoA carboxylase inhibitor
  • Another aspect embodying the present invention is (i) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor, or a long-acting conjugate thereof, and (ii) an FXR agonist, the prevention of liver disease or a pharmaceutical kit for treatment.
  • the kit is characterized in that it further comprises (iii) an ACC inhibitor.
  • Another embodiment embodying the present invention is a liver comprising the step of administering and/or using the combination, a pharmaceutical composition for the treatment or prevention of liver disease, or the pharmaceutical kit to an individual in need thereof A method for preventing or treating a disease.
  • Another aspect embodying the present invention is the use of the combination, pharmaceutical composition, or pharmaceutical kit for the prophylaxis or treatment of liver disease and/or use for the manufacture of a medicament for the prophylaxis or treatment of liver disease.
  • One aspect embodying the present invention is a pharmaceutical composition for the treatment or prevention of liver disease comprising a triple agonistic long-acting conjugate or a triple agonist, wherein the pharmaceutical composition is Provided is a pharmaceutical composition characterized in that it is used in combination with a farnesoid X receptor agonist (FXR agonist).
  • FXR agonist farnesoid X receptor agonist
  • the "triple-acting long-acting conjugate” is a glucagon receptor, GLP-1 (Glucagon-like peptide-1) receptor, and GIP (Glucose-dependent insuliontropic polypeptide) having activity on the receptor, the peptide having its in vivo half-life It refers to a long-acting conjugate to which the increasing biocompatible material part is bound.
  • the biocompatible material may be mixed with a carrier.
  • the "substance (or peptide) having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor” may be referred to as a "triple agonis, trigonal agonist” or "triple activator".
  • the "triple-acting long-acting conjugate”, which is a conjugate of the peptide, may exhibit increased durability of potency compared to the peptide to which a carrier is not bound, and in the present invention, such a conjugate is referred to as a "long-acting conjugate” and “conjugate”, “long-acting conjugate of triple activator”, or “conjugate of substances having activity against glucagon receptor, GLP-1 receptor, and GIP receptor” may be used interchangeably.
  • the pharmaceutical composition is a pharmaceutical composition for the treatment or prevention of liver disease containing a pharmaceutically effective amount of a triple-acting long-acting conjugate or a triple agent and a pharmaceutically acceptable excipient.
  • the pharmaceutical composition is characterized in that it is used in combination with a farnesoid X receptor agonist.
  • Lx is a linker containing an ethylene glycol repeating unit, and x is 0 or a natural number
  • Fc is an immunoglobulin Fc region
  • Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 1,
  • Xaa1 is histidine (His, H), 4-imidazoacetyl (CA) or tyrosine (Tyr, Y),
  • Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid or 2-aminoisobutyric acid (Aib),
  • Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q),
  • Xaa7 is threonine (Thr, T) or isoleucine (Ile, I),
  • Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C) or valine (Val, V);
  • Xaa12 is lysine (Lys, K), serine (Ser, S) or isoleucine (Ile, I);
  • Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A) or cysteine (Cys, C);
  • Xaa14 is leucine (Leu, L), methionine (Met, M) or tyrosine (Tyr, Y);
  • Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E) or leucine (Leu, L),
  • Xaa16 is glycine (Gly, G), glutamic acid (Glu, E) or serine (Ser, S);
  • Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C) or lysine (Lys, K);
  • Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R) or histidine (His, H);
  • Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C) or valine (Val, V);
  • Xaa20 is lysine (Lys, K), glutamine (Gln, Q) or arginine (Arg, R);
  • Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C) or aspartic acid (Asp, D);
  • Xaa23 is isoleucine (Ile, I) or valine (Val, V),
  • Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D) or glutamic acid (Glu, E),
  • Xaa27 is valine (Val, V), leucine (Leu, L) or lysine (Lys, K);
  • Xaa28 is cysteine (Cys, C), lysine (Lys, K), alanine (Ala, A), asparagine (Asn, N) or aspartic acid (Asp, D);
  • Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q), threonine (Thr, T), glutamic acid (Glu, E) or histidine (His, H);
  • Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K) or histidine (His, H), or is absent;
  • R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107) or m-SSGQPPPS-n (SEQ ID NO: 108), or is absent;
  • m is -Cys-, -Pro- or -Gly-Pro-;
  • n is absent or is -Cys-, -Gly-, -Ser- or -His-Gly-.
  • Xaa27 may be exceptionally methionine (Met, M).
  • the pharmaceutical composition may be used in combination with an acetyl-Coenzyme A carboxylase inhibitor (ACC inhibitor).
  • ACC inhibitor acetyl-Coenzyme A carboxylase inhibitor
  • Another aspect of the present invention is (i) a substance having activity on glucagon receptor, GLP-1 receptor, and GIP receptor or a long-acting conjugate thereof, and (ii) FXR agonist (farnesoid X receptor agonist) in combination treatment provide academic use.
  • Another embodiment includes (i) a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors or long-acting conjugates thereof, (ii) FXR agonists, and (iii) ACC inhibitors (acetyla-CoA carboxylase inhibitors) ) provides a therapeutic use in combination.
  • Another embodiment provides a pharmaceutical composition for preventing or treating liver disease using (i) a triple-acting long-acting conjugate or a triple agonist and (ii) an FXR agonist.
  • Another embodiment provides a pharmaceutical composition for the prophylaxis or treatment of liver disease using (i) a triple acting long-acting conjugate or triple agent, (ii) an FXR agonist, and (iii) an ACC inhibitor.
  • one aspect of the present invention relates to a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and a combination, pharmaceutical composition, or kit comprising an FXR agonist.
  • the triple agonist or a long-acting conjugate thereof; and the combination, pharmaceutical composition, or kit comprising a FXR agonist may be a combination, pharmaceutical composition, or kit for the prophylaxis or treatment of liver disease.
  • the combination may further comprise an ACC inhibitor, in this case, a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate) ; FXR agonists; and an ACC inhibitor.
  • an ACC inhibitor in this case, a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate) ; FXR agonists; and an ACC inhibitor.
  • the combination, pharmaceutical composition, or kit comprising an ACC inhibitor may be a combination, pharmaceutical composition, or kit for the prophylaxis or treatment of liver disease.
  • the term "combination” refers to (a) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) ); and (b) an FXR agonist, or (b') an FXR agonist and an ACC inhibitor.
  • This includes, but is not limited to, a triple-acting long-acting conjugate or a pharmaceutical composition characterized in that a triple-agonist and an FXR agonist are used in combination, or a composition in which an ACC inhibitor is additionally used in combination with the pharmaceutical composition.
  • a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (b) concomitant administration of an FXR agonist may be used interchangeably as “combined administration” or “combined administration”, wherein (a) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) ) or a conjugate thereof (triple-acting long-acting conjugate); and (b') co-administration of an FXR agonist and an ACC inhibitor may be used interchangeably as “triple combined administration” and “triple combined administration”.
  • the combination or pharmaceutical composition is
  • a) (i) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) and (ii) an FXR agonist mixed It is administered as a mixture, or (i) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting type) conjugate), (ii) an FXR agonist, and (iii) an ACC inhibitor are administered in one mixture; or
  • an FXR agonist and an ACC inhibitor are in an isolated form, glucagon receptor, GLP-1 receptor , and substances having activity on the GIP receptor (triple agonist) or long-acting conjugates thereof (triple-acting long-acting conjugates) and FXR agonists; or a substance having activity on the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist and an ACC inhibitor are formulated in separate formulations, , sequentially
  • “concomitant administration”, “combination” and “in combination” do not only mean simultaneous administration, but also a) (i) having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor. a substance (a triple agonist) or a long-acting conjugate thereof (a triple-acting long-acting conjugate) and (ii) an FXR agonist; or b) (i) a GLP-1 receptor, and a substance having activity on the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), (ii) an FXR agonist and (iii) an ACC
  • the inhibitors act together on the subject so that each of the substances ((i), (ii) and/or (iii)) performs at a level equal to or greater than its intended function.
  • composition comprising a combination refers to a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple action lasting type conjugates) and FXR agonists; or a combination comprising a GLP-1 receptor, and a substance having activity on the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist and an ACC inhibitor, or and may have therapeutic uses, but is not limited thereto. For example, it may have a use for preventing or treating liver disease, but is not limited thereto.
  • composition comprising a combination may be used interchangeably with “composition”.
  • substances having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonists) or long-acting conjugates thereof (triple-acting long-acting conjugates) and FXR agonists;
  • the GLP-1 receptor, and a substance having activity on the GIP receptor (triple agonist) or a long-acting conjugate thereof an FXR agonist and an ACC inhibitor may be administered simultaneously, separately, sequentially, or in reverse order, but is not limited thereto does not
  • the term "kit” refers to a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) and an FXR agonist; or a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist and an ACC inhibitor, It may be one comprising the combination or composition according to the invention.
  • the kit according to the present invention includes a substance having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor formulated as one formulation (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) ) and FXR agonists; or substances having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonist) or long-acting conjugates thereof (triple-acting long-acting conjugates), FXR agonists and ACC inhibitors and may additionally include a material necessary for the combined administration of two substances or three substances, but is not limited thereto.
  • the present invention relates to a substance having activity against glucagon receptors, GLP-1 receptors, and GIP receptors having a prophylactic or therapeutic effect on liver disease (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) FXR agent; Or when used in combination with an FXR agonist and an ACC inhibitor, a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting type) It was confirmed that the preventive or therapeutic effect of liver disease is dramatically improved compared to the conjugate) alone, and thus the combination therapy was provided.
  • the “substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor” may be a “peptide having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor”.
  • “triple agonist” or “triple active agent” may be used interchangeably.
  • the triple activator may be Z, which is a component of Formula 1 above.
  • Such peptides include various substances with significant levels of activity on glucagon, GLP-1, and GIP receptors, such as various peptides.
  • the triple activator having a significant level of activity on the glucagon, GLP-1, and GIP receptors includes one or more receptors of glucagon, GLP-1, and GIP receptors, specifically two or more More specifically, the in vitro activity for all three receptors is about 0.001% or more, about 0.01% or more, compared to the native ligand (natural glucagon, native GLP-1, and native GIP) of the corresponding receptor; about 0.1% or more, about 1% or more, about 2% or more, about 3% or more, about 4% or more, about 5% or more, about 6% or more, about 7% or more, about 8% or more, about 9% or more, about 10% or more, about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, about 100% or more, about It may represent 150% or more, about 200% or more, but a significantly increased range is included without limitation
  • the activity on the receptor is about 0.001% or more, 0.01% or more, 0.1% or more, 1% or more, 2% or more, 3% or more, 4% or more, 5% or more, the in vitro activity of the receptor compared to the native type. , 6% or more, 7% or more, 8% or more, 9% or more, 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, 90 % or more, 100% or more, or about 200% or more may be exemplified. However, the present invention is not limited thereto.
  • the term “about” includes all values within a range including ⁇ 0.5, ⁇ 0.4, ⁇ 0.3, ⁇ 0.2, ⁇ 0.1, etc., and includes all values in a range equal to or similar to the value following the term about, but not limited
  • Example 3 of the present specification may be referred to, but is not particularly limited thereto.
  • the peptide is characterized in that it possesses one or more, two or more, specifically three activities of the following i) to iii), specifically, it possesses a significant activity:
  • activating the receptor means that the in vitro activity of the receptor compared to the native type is about 0.001% or more, about 0.1% or more, about 1% or more, about 2% or more, about 3% or more, about 4% or more, about 5 % or more, about 6% or more, about 7% or more, about 8% or more, about 9% or more, about 10% or more, about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60 % or more, about 70% or more, about 80% or more, about 90% or more, about 100% or more, about 150% or more, about 200% or more may be exemplified. However, the present invention is not limited thereto.
  • the peptide may have an increased half-life in the body compared to any one of native GLP-1, native glucagon, and native GIP, but is not particularly limited thereto.
  • the peptide may be non-naturally occurring.
  • Such a peptide may include an intramolecular bridge (eg, a covalent bridge or a non-covalent bridge), and specifically may be in a form containing a ring, for example, at positions 16 and 20 of the peptide. It may be in the form of a ring formed between amino acids, but is not particularly limited thereto.
  • the amino acid at position 16 may be glutamic acid
  • the amino acid at position 20 may be lysine, but is not limited thereto.
  • Non-limiting examples of the ring may include a lactam bridge (or lactam ring).
  • the peptide includes all those modified to include a ring and an amino acid capable of forming a ring at a desired position.
  • amino acid pair at positions 16 and 20 of the peptide may be substituted with glutamic acid or lysine capable of forming a ring, respectively, but is not limited thereto.
  • Such a ring may be formed between amino acid side chains in the peptide, for example, a lactam ring may be formed between a lysine side chain and a glutamic acid side chain, but is not particularly limited thereto.
  • the amino acid sequence is different from that of native glucagon by one or more, and the alpha-carbon of the N-terminal amino acid residue has been removed, glucagon receptor, GLP-1 receptor, and activity against GIP receptor
  • amino acids may be substituted with other amino acids or non-natural compounds in order to avoid the recognition action of an activator degrading enzyme in order to increase the half-life in the body.
  • the peptide may be a peptide having an increased half-life in the body by avoiding the recognition action of the degrading enzyme through substitution of the second amino acid sequence among the amino acid sequence of the peptide, but amino acid substitution or change to avoid the recognition action of the degrading enzyme in the body included without limitation.
  • modifications for the production of peptides include modifications with L- or D-form amino acids, and/or non-natural amino acids; and/or by modifying the native sequence, e.g., modification of side chain functional groups, intramolecular covalent bonds, such as inter-side chain ring formation, methylation, acylation, ubiquitination, phosphorylation, aminohexylation, biotinylation, etc. includes all that
  • amino acids to be substituted or added may be atypical or non-naturally occurring amino acids as well as the 20 amino acids commonly observed in human proteins.
  • Commercial sources of atypical amino acids include Sigma-Aldrich, ChemPep and Genzyme Pharmaceuticals. Peptides containing these amino acids and canonical peptide sequences can be synthesized and purchased from commercial peptide synthesis companies, for example, American peptide company or Bachem in the United States, or Anygen in Korea.
  • Amino acid derivatives can also be obtained in the same manner, and 4-imidazoacetic acid can be used, to name just a few examples.
  • the peptide according to the present invention has its N-terminus and/or C-terminus chemically modified or protected by an organic group in order to protect it from proteolytic enzymes in vivo and to increase stability, or amino acids are added to the peptide terminus, etc. It may be added and modified form.
  • the N-terminus is acetylated and/or the C-terminus is amidated to remove these charges.
  • it is not particularly limited thereto.
  • the peptide according to the present invention includes both the peptide itself, a salt thereof (eg, a pharmaceutically acceptable salt of the peptide), or a solvate thereof.
  • the peptide may be in any pharmaceutically acceptable form.
  • the type of the salt is not particularly limited. However, it is preferable that the form is safe and effective for an individual, such as a mammal, but is not particularly limited thereto.
  • pharmaceutically acceptable means a substance that can be effectively used for a desired purpose without causing excessive toxicity, irritation, or allergic reaction within the scope of medical judgment.
  • salts derived from pharmaceutically acceptable inorganic acids, organic acids, or bases include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfonic acid, tartaric acid, acetic acid, citric acid, methanesulfonic acid, formic acid , benzoic acid, malonic acid, naphthalene-2-sulfonic acid, benzenesulfonic acid, and the like.
  • Salts derived from suitable bases may include alkali metals such as sodium and potassium, alkaline earth metals such as magnesium, and ammonium.
  • solvate refers to a compound in which the peptide or salt thereof according to the present invention forms a complex with a solvent molecule.
  • the peptide (triple agonist), or Z of Formula 1 may include an amino acid sequence represented by Formula 1 below.
  • Xaa1 is histidine, 4-imidazoacetyl, or tyrosine;
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
  • Xaa3 is glutamic acid or glutamine
  • Xaa7 is threonine or isoleucine
  • Xaa10 is leucine, tyrosine, lysine, cysteine, or valine;
  • Xaa12 is lysine, serine, or isoleucine
  • Xaa13 is glutamine, tyrosine, alanine, or cysteine;
  • Xaa14 is leucine, methionine, or tyrosine
  • Xaa15 is cysteine, aspartic acid, glutamic acid, or leucine
  • Xaa16 is glycine, glutamic acid, or serine
  • Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine;
  • Xaa18 is alanine, glutamine, arginine, or histidine;
  • Xaa19 is alanine, glutamine, cysteine, or valine;
  • Xaa20 is lysine, glutamine, or arginine
  • Xaa21 is glutamic acid, glutamine, leucine, cysteine, or aspartic acid;
  • Xaa23 is isoleucine or valine
  • Xaa24 is alanine, glutamine, cysteine, asparagine, aspartic acid, or glutamic acid,
  • Xaa27 is valine, leucine, or lysine
  • Xaa28 is cysteine, lysine, alanine, asparagine, or aspartic acid
  • Xaa29 is cysteine, glycine, glutamine, threonine, glutamic acid, or histidine;
  • Xaa30 is cysteine, glycine, lysine, or histidine, or is absent;
  • R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or is absent;
  • m is -Cys-, -Pro-, or -Gly-Pro-;
  • n is -Cys-, -Gly-, -Ser-, or -His-Gly-, or absent.
  • SEQ ID NO: 12 may be exceptionally methionine.
  • triple activator triple activator or Z of Formula 1
  • SEQ ID NOs: 1 to 11, 13 to 102 may be configured (essentially) of an amino acid sequence selected from the group consisting of, but is not limited thereto.
  • the triple active agent (triple agent) or Z of Formula 1 is SEQ ID NO: 21, 22, 42, 43, 50, 64, 66, 67, 70, 71, 76, 77, 96, It may include or (essentially) an amino acid sequence selected from the group consisting of 97 and 100, but is not limited thereto.
  • the triple active agent (triple agent) or Z of Formula 1 is selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 66, 67, 77, 96, 97 and 100 It may include or (essentially) consist of an amino acid sequence, but is not limited thereto.
  • the triple activator (triple agonist) or Z of Formula 1 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 77 and 96, or (essential ), but is not limited thereto.
  • Xaa14 may be leucine or methionine
  • Xaa15 may be cysteine, aspartic acid, or leucine.
  • a peptide examples include, but are not particularly limited to, a peptide comprising or (essentially) consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 11, 14 to 17, and 21 to 102.
  • Such a peptide may significantly activate one or more of a glucagon receptor, a GLP-1 receptor, and a GIP receptor, but is not particularly limited thereto. Specifically, it may significantly activate GLP-1 or further significantly activate glucagon receptor and/or GIP receptor, but is not particularly limited thereto.
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine, or valine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, glutamine, or cysteine
  • Xaa14 is leucine, tyrosine, or methionine
  • Xaa15 is cysteine, leucine, glutamic acid, or aspartic acid
  • Xaa17 is glutamine, arginine, isoleucine, cysteine, glutamic acid, or lysine;
  • Xaa18 is alanine, glutamine, arginine, or histidine;
  • Xaa19 is alanine, glutamine, valine, or cysteine
  • Xaa20 is lysine, arginine, or glutamine
  • Xaa21 is glutamic acid, glutamine, leucine, cysteine, or aspartic acid;
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid, or aspartic acid;
  • Xaa27 may be a peptide, which is leucine or lysine, but is not particularly limited thereto.
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine, or valine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, or cysteine
  • Xaa14 is leucine or methionine
  • Xaa15 is cysteine or aspartic acid
  • Xaa17 is glutamine, arginine, isoleucine, cysteine, or lysine;
  • Xaa18 is alanine, arginine, or histidine
  • Xaa19 is alanine, glutamine, or cysteine
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid, cysteine, or aspartic acid
  • Xaa23 is valine
  • Xaa24 is alanine, glutamine, cysteine, asparagine, or aspartic acid
  • Xaa27 may be leucine or lysine, but is not particularly limited thereto.
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa7 is threonine
  • Xaa10 is tyrosine or cysteine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, or cysteine
  • Xaa14 is leucine or methionine
  • Xaa15 is cysteine or aspartic acid
  • Xaa16 is glutamic acid
  • Xaa17 is arginine, isoleucine, cysteine, or lysine;
  • Xaa18 is alanine, arginine, or histidine
  • Xaa19 is alanine, glutamine, or cysteine
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid or aspartic acid
  • Xaa23 is valine
  • Xaa24 is glutamine, asparagine, or aspartic acid
  • Xaa27 is leucine
  • Xaa28 may be cysteine, alanine, asparagine, or aspartic acid.
  • Xaa1 is histidine or 4-imidazoacetyl
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa3 is glutamine
  • Xaa7 is threonine
  • Xaa10 is tyrosine
  • Xaa12 is isoleucine
  • Xaa13 is alanine or cysteine
  • Xaa14 is methionine
  • Xaa15 is aspartic acid
  • Xaa16 is glutamic acid
  • Xaa17 is isoleucine or lysine
  • Xaa18 is alanine or histidine
  • Xaa19 is glutamine or cysteine
  • Xaa20 is lysine
  • Xaa21 is aspartic acid
  • Xaa23 is valine
  • Xaa24 is asparagine
  • Xaa27 is leucine
  • Xaa28 is alanine or asparagine
  • Xaa29 is glutamine or threonine
  • Xaa30 may be cysteine or lysine or absent.
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
  • Xaa3 is glutamine
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine, or valine
  • Xaa12 is lysine
  • Xaa13 is tyrosine
  • Xaa14 is leucine
  • Xaa15 is aspartic acid
  • Xaa16 is glycine, glutamic acid, or serine
  • Xaa17 is glutamine, arginine, cysteine, or lysine
  • Xaa18 is alanine, arginine, or histidine
  • Xaa19 is alanine or glutamine
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid, cysteine, or aspartic acid
  • Xaa23 is valine
  • Xaa24 is alanine, glutamine, or cysteine
  • Xaa27 is leucine or lysine
  • Xaa29 may be glycine, glutamine, threonine, or histidine, but is not particularly limited thereto.
  • These peptides have significant activation levels of GLP-1 receptors and glucagon receptors, and are higher than those of GIP receptors;
  • the activation levels of the GLP-1 receptor, the glucagon receptor and the GIP receptor are all significant;
  • the degree of activation of the GLP-1 receptor and the GIP receptor is significant and may correspond to a case where the activation level of the glucagon receptor is higher than that of the glucagon receptor, but is not particularly limited thereto.
  • Such peptides include SEQ ID NOs: 8, 9, 21 to 37, 39, 42, 43, 49 to 61, 64 to 83, 85, 86, 88, 89, 91 to 93, selected from the group consisting of 95 to 102 and a peptide comprising or (essentially) consisting of an amino acid sequence, but is not particularly limited thereto.
  • the peptide may include an amino acid sequence represented by the following general formula (2).
  • Xaa1 is 4-imidazoacetyl, histidine, or tyrosine;
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
  • Xaa10 is tyrosine or cysteine
  • Xaa13 is alanine, glutamine, tyrosine, or cysteine;
  • Xaa14 is leucine, methionine, or tyrosine
  • Xaa15 is aspartic acid, glutamic acid, or leucine
  • Xaa16 is glycine, glutamic acid, or serine
  • Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine;
  • Xaa18 is alanine, glutamine, arginine, or histidine;
  • Xaa19 is alanine, glutamine, cysteine, or valine;
  • Xaa20 is lysine, glutamine, or arginine
  • Xaa21 is cysteine, glutamic acid, glutamine, leucine, or aspartic acid;
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, alanine, glutamine, asparagine, or glutamic acid
  • Xaa28 is lysine, cysteine, asparagine, or aspartic acid
  • Xaa29 is glycine, glutamine, cysteine, or histidine
  • Xaa30 is cysteine, glycine, lysine, or histidine
  • Xaa31 is proline or cysteine
  • Xaa40 is cysteine or absent.
  • Xaa13 is alanine, tyrosine, or cysteine
  • Xaa15 is aspartic acid or glutamic acid
  • Xaa17 is glutamine, arginine, cysteine, or lysine
  • Xaa18 is alanine, arginine, or histidine
  • Xaa21 is cysteine, glutamic acid, glutamine, or aspartic acid
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, glutamine, or asparagine
  • Xaa28 is cysteine, asparagine, or aspartic acid
  • Xaa29 is glutamine, cysteine, or histidine
  • Xaa30 may be cysteine, lysine, or histidine.
  • Examples of such a peptide include an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 64 to 77, and 95 to 102, or SEQ ID NOs: 21, 22, 42, 43, 50, 64 to 77, and a peptide comprising or (essentially) consisting of an amino acid sequence selected from the group consisting of 96 to 102, but is not particularly limited thereto.
  • the peptide may include an amino acid sequence of the following general formula (3).
  • Xaa1 is histidine or tyrosine
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa13 is alanine, tyrosine or cysteine
  • Xaa17 is arginine, cysteine, or lysine
  • Xaa18 is alanine or arginine
  • Xaa19 is alanine or cysteine
  • Xaa21 is glutamic acid or aspartic acid
  • Xaa24 is glutamine or asparagine
  • Xaa28 is cysteine or aspartic acid
  • Xaa29 is cysteine, histidine, or glutamine
  • Xaa30 is cysteine or histidine
  • Xaa31 is proline or cysteine
  • Xaa40 may be cysteine or absent.
  • peptides examples include peptides comprising or (essentially) consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 64 to 71, 75 to 77, and 96 to 102. However, it is not particularly limited thereto.
  • R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), CSSGQPPPS (SEQ ID NO: 109), GPSSGAPPPS (SEQ ID NO: 110), GPSSGAPPPSC (SEQ ID NO: 111), PSSGAPPPS (SEQ ID NO: 112), PSSGAPPPSG (SEQ ID NO: SEQ ID NO: 110) 113), PSSGAPPPSHG (SEQ ID NO: 114), PSSGAPPPSS (SEQ ID NO: 115), PSSGQPPPS (SEQ ID NO: 116), or PSSGQPPPSC (SEQ ID NO: 117), or absent, but is not particularly limited thereto.
  • the length of the peptide of the present invention can be synthesized by a method well known in the art, for example, an automatic peptide synthesizer, or it can be produced by a genetic engineering technique.
  • the peptides of the present invention can be prepared by standard synthetic methods, recombinant expression systems, or any other method in the art.
  • the peptides according to the invention can be synthesized in a number of ways, including, for example, those comprising:
  • a method for obtaining a fragment of a peptide by any combination of (a), (b) and (c), and then ligating the fragments to obtain a peptide, and recovering the peptide.
  • the triple-acting long-acting binder or the triple-acting agent may be non-naturally occurring.
  • the triple-functional long-acting conjugate may be a conjugate represented by Formula 1 described above, but is not limited thereto.
  • Lx is a linker containing an ethylene glycol repeating unit, and x is 0 or a natural number
  • Fc is an immunoglobulin Fc region
  • Z is as described above. Z may mean triple agonist.
  • Fc is a substance capable of increasing the half-life of a peptide, having activity against Z, that is, glucagon receptor, GLP-1 receptor, and GIP receptor, in one component of the moiety constituting the conjugate of the present invention. corresponds to
  • the Fc may be bonded to each other by a covalent chemical bond or a non-covalent chemical bond with Z, and Fc and Z may be bonded to each other through Lx by a covalent chemical bond, a non-covalent chemical bond, or a combination thereof.
  • the Fc may be directly linked to Z (ie, x is 0 in Formula 1) or linked through a linker (Lx).
  • Lx may be a non-peptidyl linker, for example, a linker containing an ethylene glycol repeating unit.
  • non-peptidyl linker includes a biocompatible polymer in which two or more repeating units are bonded. The repeating units are linked to each other through any covalent bond other than a peptide bond.
  • the non-peptidyl linker may be one component constituting a moiety of the conjugate of the present invention, and corresponds to Lx in Formula 1 above.
  • x may be 1 or more, and when x is 2 or more, each L may be independent.
  • the non-peptidyl linker that can be used in the present invention may be used without limitation as long as it is a polymer resistant to proteolytic enzymes in vivo.
  • the non-peptidyl linker may be used in combination with a non-peptidyl polymer.
  • non-peptidyl linker may be a linker containing an ethylene glycol repeating unit, for example, polyethylene glycol, and also derivatives thereof known in the art and easily at the level of skill in the art. Derivatives that can be prepared are also included in the scope of the present invention.
  • the repeating unit of the non-peptidyl linker may be an ethylene glycol repeating unit, and specifically, the non-peptidyl linker may include an ethylene glycol repeating unit and a functional group used in the preparation of the conjugate at the terminal thereof.
  • the long-acting conjugate according to the present invention may be in a form in which Z and Fc are linked through the functional group, but is not limited thereto.
  • the non-peptidyl linker may include two, or three or more functional groups, and each functional group may be the same or different from each other, but is not limited thereto.
  • the linker may be polyethylene glycol (PEG) represented by the following formula (2), but is not limited thereto:
  • the PEG moiety in the long-acting conjugate may include, but is not limited to, the -(CH2CH2O)n- structure as well as an oxygen atom intervening between the linking element and the -(CH2CH2O)n-.
  • the conjugate comprises a peptide (Z) comprising the amino acid sequence of Formula 1 or an amino acid sequence of any one of SEQ ID NOs: 1 to 102 and an immunoglobulin Fc region (Fc) comprising an ethylene glycol repeating unit It may be a structure connected by a covalent bond through a linker containing it, but is not limited thereto.
  • the polyethylene glycol is a term encompassing all forms of ethylene glycol homopolymer, PEG copolymer, or monomethyl-substituted PEG polymer (mPEG), but is not particularly limited thereto.
  • the ethylene glycol repeating unit may be represented by, for example, [OCH 2 CH 2 ]n, and the value of n is a natural number, the average molecular weight of the [OCH 2 CH 2 ]n site in the peptide conjugate, such as the number
  • the average molecular weight may be set to be greater than 0 to about 100 kDa, but is not limited thereto.
  • the n value is a natural number
  • the average molecular weight of the [OCH 2 CH 2 ]n site in the peptide conjugate for example, has a number average molecular weight of about 1 to about 100 kDa, about 1 to about 80 kDa, about 1 to about 50 kDa, about 1 to about 30 kDa, about 1 to about 25 kDa, about 1 to about 20 kDa, about 1 to about 15 kDa, about 1 to about 13 kDa, about 1 to about 11 kDa, about 1 to about 10 kDa, about 1 to about 8 kDa, about 1 to about 5 kDa, about 1 to about 3.4 kDa, about 3 to about 30 kDa, about 3 to about 27 kDa, about 3 to about 25 kDa, about 3 to about 22 kDa , about 3 to about 20 kDa, about 3 to about 18 kDa, about 3 to about 16 kDa
  • the non-peptidyl linker that can be used in the present invention may be used without limitation as long as it is a polymer including an ethylene glycol repeating unit resistant to proteolytic enzymes in vivo.
  • the molecular weight of the non-peptidyl polymer is, but is not limited to, greater than 0, in the range of about 100 kDa, in the range of about 1 to about 100 kDa, specifically in the range of about 1 to about 20 kDa, or in the range of about 1 to about 10 kDa.
  • a combination of different types of polymers as well as one type of polymer may be used for the non-peptidyl linker of the present invention that is coupled to the polypeptide corresponding to the Fc.
  • both ends of the non-peptidyl linker are each Fc, such as an amine group, a thiol group, a hydroxyl group, and an amine group, a thiol group, an azide group, and a hydroxyl group of an immunoglobulin Fc region. It may be combined, but is not limited thereto.
  • the non-peptidyl polymer has a reactive group capable of binding to Fc (eg, immunoglobulin Fc region) and Z, respectively, at both ends, specifically, a thiol group of a cysteine in the immunoglobulin Fc region; an amine group located at the N-terminus, lysine, arginine, glutamine and/or histidine; and/or bonded to a hydroxyl group located at the C-terminus, and a thiol group of cysteine of Z; amine groups of lysine, arginine, glutamine and/or histidine; azide group of azidolysin; and/or a reactive group capable of bonding to a hydroxyl group, but is not limited thereto.
  • Fc immunoglobulin Fc region
  • Z a reactive group capable of binding to Fc (eg, immunoglobulin Fc region) and Z, respectively, at both ends, specifically, a thiol group of a cysteine in the immuno
  • both ends of the non-peptidyl linker may be respectively bound to an amine or thiol group of an Fc region, such as an immunoglobulin Fc region, and an amine or thiol group of Z.
  • the non-peptidyl polymer has a reactive group capable of binding to Fc (eg, immunoglobulin Fc region) and Z, respectively, at both ends, specifically Z, or the N-terminus of Fc (eg, immunoglobulin Fc region).
  • Fc eg, immunoglobulin Fc region
  • Z the N-terminus of Fc
  • it may include a reactive group capable of bonding to an amine group located on lysine, or a thiol group of cysteine, but is not limited thereto.
  • the reactive group of the non-peptide polymer capable of binding to Fc may be at least one selected from the group consisting of an aldehyde group, a maleimide group and a succinimide derivative, but is not limited thereto. .
  • the aldehyde group may be exemplified by a propionaldehyde group or a butyl aldehyde group, but is not limited thereto.
  • succinimidyl valerate succinimidyl methylbutanoate, succinimidyl methylpropionate, succinimidyl butanoate, succinimidyl propionate, N-hydroxysuccini Mead, hydroxy succinimidyl, succinimidyl carboxymethyl or succinimidyl carbonate may be used, but are not limited thereto.
  • the non-peptide linker may be connected to Z and Fc through such a reactive group and converted into a non-peptide linker linkage, but is not particularly limited thereto.
  • the final product resulting from reductive amination by aldehyde bonds is much more stable than those linked by amide bonds.
  • the aldehyde reactive group selectively reacts with the N-terminus at a low pH, and can form a covalent bond with a lysine residue at a high pH, for example, pH 9.0.
  • the reactive groups at both ends of the non-peptidyl linker may be the same or different from each other, for example, a maleimide group at one end and an aldehyde group, a propionaldehyde group, or a butyl aldehyde group at the other end.
  • Fc specifically, immunoglobulin Fc region and Z can be bound to each end of the non-peptidyl linker, it is not particularly limited thereto.
  • one end of the non-peptidyl linker may include a maleimide group as a reactive group, and an aldehyde group, a propionaldehyde group, or a butyl aldehyde group at the other end of the non-peptidyl linker.
  • the hydroxyl group can be activated into the various reactive groups by a known chemical reaction, or a commercially available polyethylene glycol having a modified reactive group is used.
  • the long-acting protein conjugate of the invention can be prepared.
  • the non-peptidyl polymer may be linked to a cysteine residue of Z, more specifically, a -SH group of cysteine, but is not limited thereto.
  • cysteine residue 10 cysteine 13, cysteine 15, cysteine 17, cysteine 19, cysteine 21, cysteine 24, cysteine 28, 29
  • the non-peptidyl polymer may be linked to cysteine residue No. 30, cysteine residue 30, cysteine 31, cysteine 40, or cysteine 41, but is not particularly limited thereto.
  • a reactive group of the non-peptidyl polymer may be linked to the -SH group of the cysteine residue, and all of the above descriptions apply to the reactive group.
  • maleimide-PEG-aldehyde is used, the maleimide group is linked to the -SH group of Z by a thioether bond, and the aldehyde group is Fc, specifically, the -NH 2 group of immunoglobulin Fc reductive amination reaction may be connected through, but is not limited thereto, and this corresponds to one example.
  • the N-terminal amino group of the immunoglobulin Fc region is linked to an oxygen atom located at one end of PEG, a non-peptidyl polymer, through a linker functional group having the structure -CH 2 CH 2 CH 2 - , -PEG-O-CH 2 CH 2 CH 2 NH-Can form the same structure as immunoglobulin Fc, and one end of PEG through a thioether bond is a sequence of any one of Formula 1 or SEQ ID NOs: 1 to 102 It can form a structure linked to the sulfur atom located at the cysteine of the peptide containing.
  • the above-mentioned thioether bond is may contain the structure of
  • the non-peptidyl polymer may be linked to a lysine residue of Z, more specifically, an amino group of lysine, but is not limited thereto.
  • the reactive group of the non-peptidyl polymer may be linked to -NH 2 located at the N-terminus of the immunoglobulin Fc region, but this corresponds to one example.
  • the peptide may be linked to a linker having a reactive group through the C-terminus, but this corresponds to one example.
  • C-terminus refers to the carboxy terminus of a peptide, and refers to a position capable of binding to a linker for the purpose of the present invention.
  • it may include all amino acid residues around the C-terminus as well as the most terminal amino acid residue at the C-terminus, and specifically includes the first to 20th amino acid residues from the most terminal. can, but is not limited thereto.
  • the Fc may be an immunoglobulin Fc region, and more specifically, the immunoglobulin Fc region may be derived from IgG, but is not particularly limited thereto.
  • immunoglobulin Fc region refers to a region including heavy chain constant region 2 (CH2) and/or heavy chain constant region 3 (CH3), excluding the heavy and light chain variable regions of immunoglobulin.
  • the immunoglobulin Fc region may be one component constituting a moiety of the conjugate of the present invention.
  • the immunoglobulin Fc region may be used interchangeably with “immunoglobulin Fc fragment”.
  • the Fc region not only the native sequence obtained from papain digestion of immunoglobulin, but also its derivatives, such as one or more amino acid residues in the native sequence, are converted by deletion, insertion, non-conservative or conservative substitution, or a combination thereof, resulting in a native natural sequence. It encompasses and includes sequences that differ from the type.
  • the Fc has a structure in which two polypeptide chains are linked by a disulfide bond, and may have a structure in which only one of the two chains is linked through a nitrogen atom, but is not limited thereto.
  • the linkage via the nitrogen atom may be linked via reductive amination to the epsilon amino atom or the N-terminal amino group of lysine.
  • Reductive amination reaction refers to a reaction in which an amine group or an amino group of a reactant reacts with an aldehyde (that is, a functional group capable of reductive amination) of another reactant to form an amine, and then forms an amine bond by a reduction reaction, It is an organic synthesis reaction well known in the art.
  • the Fc may be linked through the nitrogen atom of its N-terminal proline, but is not limited thereto.
  • the immunoglobulin Fc region is one component constituting a moiety of the conjugate of Formula 1 of the present invention, and specifically, may correspond to Fc in Formula 1 above.
  • the immunoglobulin Fc region may include a hinge region in the heavy chain constant region, but is not limited thereto.
  • the immunoglobulin Fc region may include a specific hinge sequence at the N-terminus.
  • flankinge sequence refers to a region that is located on a heavy chain and forms a dimer of an immunoglobulin Fc region through an inter disulfide bond.
  • the hinge sequence may be mutated to have only one cysteine residue by deleting a portion of the hinge sequence having the following amino acid sequence, but is not limited thereto:
  • the hinge sequence may include only one cysteine residue by deleting the 8th or 11th cysteine residue in the hinge sequence of SEQ ID NO: 118.
  • the hinge sequence of the present invention may be composed of 3 to 12 amino acids, including only one cysteine residue, but is not limited thereto.
  • the hinge sequence of the present invention may have the following sequences: Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Pro-Ser-Cys-Pro (SEQ ID NO: 119), Glu-Ser- Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser-Pro (SEQ ID NO: 120), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser (SEQ ID NO: 121), Glu- Ser-Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Pro (SEQ ID NO: 122), Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser (SEQ ID NO: 123), Glu-Ser-Lys- Tyr-Gly-Pro-Pro-Cys (SEQ ID NO: 124), Glu-Lys-Tyr-Gly-Pro-Pro-Cys (SEQ ID NO
  • the hinge sequence may include the amino acid sequence of SEQ ID NO: 128 (Pro-Ser-Cys-Pro) or SEQ ID NO: 137 (Ser-Cys-Pro), but is not limited thereto.
  • the immunoglobulin Fc region of the present invention may be in a form in which two immunoglobulin Fc chain molecules form a dimer due to the presence of a hinge sequence.
  • one end of the linker is a dimer immunoglobulin It may be in a form linked to one chain of the Fc region, but is not limited thereto.
  • N-terminus refers to the amino terminus of a protein or polypeptide, and 1, 2, 3, 4, 5, 6, It may include up to 7, 8, 9, or 10 or more amino acids.
  • the immunoglobulin Fc region of the present invention may include a hinge sequence at the N-terminus, but is not limited thereto.
  • part or all of the heavy chain constant region 1 (CH1) and/or the light chain constant region except for only the heavy and light chain variable regions of immunoglobulin 1 (CL1) may be an extended Fc region. Also, it may be a region in which some fairly long amino acid sequences corresponding to CH2 and/or CH3 have been removed.
  • the immunoglobulin Fc region of the present invention comprises 1) a CH1 domain, a CH2 domain, a CH3 domain and a CH4 domain, 2) a CH1 domain and a CH2 domain, 3) a CH1 domain and a CH3 domain, 4) a CH2 domain and a CH3 domain, 5) a combination of one or two or more domains of CH1 domain, CH2 domain, CH3 domain and CH4 domain with an immunoglobulin hinge region (or a part of the hinge region), 6) heavy chain constant region Each domain may be a dimer of a light chain constant region .
  • the present invention is not limited thereto.
  • the immunoglobulin Fc region may be in a dimeric form, and one molecule of Z may be covalently linked to one Fc region in a dimeric form, in which case the immunoglobulin Fc and Z may be covalently linked to each other through a linker containing an ethylene glycol repeating unit.
  • the immunoglobulin Fc and Z may be linked to each other by a linker containing an ethylene glycol repeating unit.
  • Z may be covalently linked via a linker Lx to only one of the two polypeptide chains of the Fc region dimer Fc.
  • Lx linker
  • the Fc region dimer Fc only one molecule of Z may be covalently linked via Lx to one polypeptide chain to which Z is linked.
  • the Fc may be a homodimer.
  • the immunoglobulin Fc region Fc is a dimer consisting of two polypeptide chains, and one end of Lx may be connected to only one of the two polypeptide chains, but is not limited thereto. does not
  • the immunoglobulin Fc region of the present invention includes a native amino acid sequence as well as a sequence derivative thereof.
  • An amino acid sequence derivative means that one or more amino acid residues in a natural amino acid sequence have a different sequence by deletion, insertion, non-conservative or conservative substitution, or a combination thereof.
  • amino acid residues 214 to 238, 297 to 299, 318 to 322, or 327 to 331 known to be important for binding may be used as suitable sites for modification.
  • various types of derivatives are possible, such as a site capable of forming a disulfide bond is removed, some amino acids at the N-terminus of native Fc are removed, or a methionine residue may be added to the N-terminus of native Fc do.
  • the complement binding site eg, the C1q binding site
  • the ADCC antibody dependent cell mediated cytotoxicity
  • the above-described Fc derivative may exhibit biological activity equivalent to that of the Fc region of the present invention, and may have increased structural stability to heat, pH, etc. of the Fc region.
  • the Fc region may be obtained from a native type isolated in vivo from animals such as humans, cows, goats, pigs, mice, rabbits, hamsters, rats or guinea pigs, or obtained from transformed animal cells or microorganisms. It may be recombinant or a derivative thereof.
  • the method of obtaining from the native type may be a method of obtaining whole immunoglobulin by isolating it from a living body of a human or animal and then treating it with a proteolytic enzyme. When treated with papain, it is cleaved into Fab and Fc, and when treated with pepsin, it is cleaved into pF'c and F(ab) 2 .
  • Fc or pF'c may be separated using size-exclusion chromatography or the like.
  • it is a recombinant immunoglobulin Fc region obtained by obtaining a human-derived Fc region from a microorganism.
  • the immunoglobulin Fc region may have a native sugar chain, an increased sugar chain compared to the native type, a decreased sugar chain compared to the native type, or a form in which the sugar chain is removed.
  • Conventional methods such as chemical methods, enzymatic methods, and genetic engineering methods using microorganisms may be used for the increase or decrease or removal of such immunoglobulin Fc sugar chains.
  • the immunoglobulin Fc region from which the sugar chains are removed from the Fc has significantly reduced binding to complement (c1q) and reduced or eliminated antibody-dependent cytotoxicity or complement-dependent cytotoxicity, so that unnecessary immune responses in vivo are not induced. does not In this respect, a form more suitable for the original purpose as a drug carrier will be an immunoglobulin Fc region in which sugar chains are removed or non-glycosylated.
  • deglycosylation refers to an Fc region from which sugars have been removed with an enzyme
  • aglycosylation refers to an Fc region that is not glycosylated by production in prokaryotes, in a more specific embodiment, in E. coli. .
  • the immunoglobulin Fc region may be of human or animal origin, such as cattle, goats, pigs, mice, rabbits, hamsters, rats, and guinea pigs, and in a more specific embodiment, it is of human origin.
  • the immunoglobulin Fc region may be an Fc region derived from IgG, IgA, IgD, IgE, or IgM, or a combination or hybrid thereof. In a more specific embodiment, it is derived from IgG or IgM, which is most abundant in human blood, and in a more specific embodiment, it is derived from IgG, which is known to enhance the half-life of ligand binding proteins. In an even more specific embodiment, the immunoglobulin Fc region is an IgG4 Fc region, and in the most specific embodiment, the immunoglobulin Fc region is a non-glycosylated Fc region derived from human IgG4, but is not limited thereto.
  • the immunoglobulin Fc fragment is a fragment of human IgG4 Fc, and is a homologous type in which two monomers are linked through a disulfide bond (inter-chain form) between cysteine, amino acid 3 of each monomer. It may be in the form of a dimer, wherein each monomer of the homodimer is independently an internal disulfide bond between cysteines at positions 35 and 95 and an internal disulfide bond between cysteines at positions 141 and 199, that is, two internal It may have/have disulfide bonds (intra-chain form).
  • the number of amino acids of each monomer may consist of 221 amino acids, and the amino acids forming the homodimer may consist of a total of 442 amino acids, but is not limited thereto.
  • two monomers having the amino acid sequence of SEQ ID NO: 138 (consisting of 221 amino acids) form a homodimer through a disulfide bond between cysteine, the 3rd amino acid of each monomer, and the homodimer
  • the monomers of may each independently form an internal disulfide bond between cysteines at positions 35 and 95 and an internal disulfide bond between cysteines at positions 141 and 199, but is not limited thereto.
  • the Fc of Formula 1 may include a monomer having the amino acid sequence of SEQ ID NO: 138, and the Fc may be a homodimer of the monomer having the amino acid sequence of SEQ ID NO: 138, but is not limited thereto.
  • the immunoglobulin Fc region may be a homodimer comprising the amino acid sequence of SEQ ID NO: 139 (consisting of 442 amino acids), but is not limited thereto.
  • the term "combination" with respect to an immunoglobulin Fc region means that when a dimer or multimer is formed, a polypeptide encoding a single-chain immunoglobulin Fc region of the same origin binds to a single-chain polypeptide of a different origin. means to form That is, it is possible to prepare a dimer or multimer from two or more fragments selected from the group consisting of IgG Fc, IgA Fc, IgM Fc, IgD Fc and IgE Fc fragment.
  • hybrid is a term meaning that sequences corresponding to immunoglobulin Fc fragments of two or more different origins exist in a single-chain immunoglobulin constant region.
  • various types of hybrids are possible. That is, a hybrid of domains consisting of 1 to 4 domains from the group consisting of CH1, CH2, CH3 and CH4 of IgG Fc, IgM Fc, IgA Fc, IgE Fc and IgD Fc is possible, and may include a hinge.
  • IgG can also be divided into subclasses of IgG1, IgG2, IgG3 and IgG4, and in the present invention, a combination thereof or hybridization thereof is also possible. Specifically, they are subclasses of IgG2 and IgG4, and most specifically, an Fc fragment of IgG4 having little effector function such as complement dependent cytotoxicity (CDC).
  • CDC complement dependent cytotoxicity
  • conjugate may have an increased duration of effect compared to native GLP-1, GIP, or glucagon, or compared to Z in which Fc is not modified. It includes all types of particles and the like, but is not limited thereto.
  • the FXR agonist refers to an agonist of FXR, a nuclear receptor activated by bile acids, also known as Bile acid Receptor (BAR), for the glucagon receptor, GLP-1 receptor, and GIP receptor of the present invention.
  • BAR Bile acid Receptor
  • Substances having an activity (triple agonist) or long-acting combination thereof and used in combination therapy to improve the effect of preventing or treating liver disease compared to alone may be included without limitation.
  • FXR is expressed at key sites of bile acid metabolism, such as the liver, intestine, and kidney, and it affects several metabolic pathways, including bile acids, in a tissue-specific manner.
  • FXR is known to inhibit bile acid production and promote elimination through various mechanisms in the liver and intestine.
  • FXR agonists reduce hepatic triglyceride synthesis to reduce steatosis, inhibit hepatic stellate cell activation to reduce hepatic fibrosis, stimulate FGF15/FGF19 expression (a major regulator of bile acid metabolism), and thereby reduce hepatic insulin sensitivity.
  • the FXR agonist is used in combination therapy together with a substance having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) of the present invention.
  • Substances capable of improving the effect of preventing or treating liver disease compared to alone are included without limitation, for example, cafestol, chenodeoxycholic acid, obeticholic acid, fexara Fexaramine, GW 4064, PX104, 6E-CDCA (6-ethyl-chedeoxycholic acid), AKN-083 or Tropifexor, Cilofexor, EDP-305, AGN-242266, AGN-242256, EP-024297, RDX-023, BWL-200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, It may be PX20606, EYP-001, TERN-101, TC-100, or INT-2228.
  • the ACC inhibitor refers to a substance that inhibits acetyla-CoA carboxylase, an enzyme important for the regulation of fatty acid production and metabolism, and is active on the glucagon receptor, GLP-1 receptor, and GIP receptor of the present invention
  • a substance having (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) and a substance that can be used in combination therapy with the FXR agonist to prevent or improve the therapeutic effect of liver disease compared to alone may be included without limitation.
  • the ACC inhibitor is a substance having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor of the present invention (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), and in combination with an FXR agonist
  • Substances that can be used for therapy and improve the effect of preventing or treating liver disease compared to alone are included without limitation, for example, CP-640186, (4-piperidinyl)-piperazine derivative ((4-piperidinyl) -piperazine derivatives), 1,4-disubstituted cyclohexane derivatives, spirochromanone derivatives, spirolactam derivatives, spirodiamine derivatives, spiropenta Thiophene pyrimidone derivatives including Spiropentacylamide derivatives, Pseudopeptide pyrrolidinedione derivatives, Macrocyclic polyketone derivatives, firsocostat derivatives), amino-
  • the combination, pharmaceutical composition or kit of the present invention can be used for the prevention or treatment of liver disease.
  • prevention refers to (a) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate) and an FXR agonist; or (b) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate), a combination comprising an FXR agonist and an ACC inhibitor; or It refers to any action that inhibits or delays the onset of a desired disease, such as liver disease, by administration of the composition, and "treatment” refers to (a) a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (3 heavy agonist) or a conjugate thereof (a triple acting long-acting conjugate) and an FXR agonist; or (b) a substance having activity on the glucagon receptor, the GIP receptor (triple agonist) or
  • the term "administration" means introducing a predetermined substance to a patient by any suitable method, and the route of administration of the composition is not particularly limited thereto, but any general route through which the composition can reach an in vivo target It can be administered through, for example, intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intradermal administration, oral administration, topical administration, intranasal administration, intrapulmonary administration, rectal administration, etc. can be .
  • liver disease refers to a disease occurring in the liver, and may include metabolic liver disease or liver inflammation, but is not limited thereto.
  • Representative examples of the liver disease include non-alcoholic fatty liver disease or cholestasis liver disease.
  • various liver diseases that cause abnormalities in the tissues and functions of the liver are within the scope of the liver disease of the present invention. can be entered, but is not limited thereto.
  • nonalcoholic fatty liver disease refers to a case in which there is no history of alcohol intake or is accompanied by fatty liver although it is not related to alcohol intake.
  • Fatty liver refers to a phenomenon in which triglycerides are abnormally deposited in liver cells, unlike normal cases. About 5% of a normal liver is composed of adipose tissue, and triglycerides, fatty acids, phospholipids, cholesterol, and cholesterol esters are the main components of fat. If it is more than 5% of the liver weight, it is diagnosed as fatty liver. Fatty liver is caused by disorders in fat metabolism in hepatocytes or defects in the process of transporting excess fat, and mainly occurs due to disorders in fat metabolism in the liver.
  • composition according to the present invention inhibits NAS (NAFLD activity score) reduction and fibrosis, it can be seen that lesions of various diseases included in the nonalcoholic fatty liver disease can be prevented or treated.
  • NAS NAFLD activity score
  • the nonalcoholic fatty liver disease includes simple steatosis, liver inflammation, nonalcoholic fatty liver, nonalcoholic steatohepatitis, cirrhosis, liver fibrosis, liver failure or liver cancer. included without
  • the nonalcoholic fatty liver disease may be at least one disease selected from the group consisting of liver inflammation, nonalcoholic steatohepatitis and liver fibrosis, but the nonalcoholic fatty liver disease prevented or treated with the composition of the present invention is included without limitation.
  • nonalcoholic steatohepatitis is one of nonalcoholic fatty liver disease, and is a representative example of liver disease accompanied by liver cell necrosis, inflammation, and fibrosis.
  • the composition according to the present invention contains NAS (NAFLD activity) score) reduction and suppression of fibrosis to exhibit an effect on nonalcoholic steatohepatitis, specifically, nonalcoholic steatohepatitis with fatty liver, liver fibrosis or cirrhosis; or liver cancer caused by nonalcoholic steatohepatitis
  • NAS NAFLD activity
  • the present invention is not limited thereto.
  • liver inflamation refers to a disease that causes inflammation of the liver as the biggest cause of liver disease, and is divided into acute hepatitis and chronic hepatitis according to the cause and symptoms. Viruses, alcohol, drugs, immune abnormalities, and metabolic diseases are the main causes.
  • liver fibrosis is a result of the wound healing process for repeated liver damage, and unlike liver cirrhosis, it is reversible, consists of thin fibrils, Normal recovery may be possible when the cause of liver damage is eliminated. progresses to cirrhosis of the liver.
  • the composition of the present invention may exhibit a preventive or therapeutic effect on liver fibrosis by reducing the hydroxyproline content in the subject to which the pharmaceutical composition is administered, but is not limited thereto.
  • the conjugate according to the present invention not only has the effect of alleviating the fibrosis of the liver itself, but also can exhibit an effect on diseases accompanying or caused by fibrosis of the liver.
  • liver cirrhosis of the present invention is a chronic disease that occurs while repeating hepatocyte regeneration and increase in fibrous tissue and is pathologically accompanied by necrosis, inflammation and fibrosis, and ultimately liver failure It progresses to liver cirrhosis complications and diseases such as liver cancer, leading to death.
  • liver cirrhosis complications and diseases such as liver cancer, leading to death.
  • it since there are no subjective symptoms in the early stage and it is discovered only at a fairly advanced stage, it is required to rapidly treat liver fibrosis, which is a condition before it evolves into cirrhosis or the like.
  • liver decompensation is due to liver damage or liver disease such as viral hepatitis, cirrhosis, drugs or alcohol, liver function is weakened, and the liver can perform protein synthesis and metabolic functions as normal physiological functions. means no state. It is divided into acute liver failure or chronic liver failure according to the rate of progression, and is known to cause various complications.
  • liver damage or liver disease such as viral hepatitis, cirrhosis, drugs or alcohol
  • liver function is weakened, and the liver can perform protein synthesis and metabolic functions as normal physiological functions. means no state. It is divided into acute liver failure or chronic liver failure according to the rate of progression, and is known to cause various complications.
  • Hepatocellular carcinoma of the present invention means a malignant tumor originating from liver cells, and can be divided into primary liver cancer (hepatocellular carcinoma) arising from hepatocytes itself and metastatic liver cancer in which cancers of other tissues have metastasized to the liver. More than 90% of liver cancers are primary liver cancers. In addition to hepatitis and chronic liver disease, the main causes are alcohol, smoking, and obesity.
  • cholestasis is a condition in which the flow of bile from the liver to the duodenum is slowed or blocked
  • cholestasis liver disease is It is meant to be hampered by conditions such as disease, extended jugular vein nutrition, or side effects of certain medications (eg, some antibiotics). Common signs of cholestasis include fatigue, pruritus (itching), jaundice, and xanthomas (deposition of high cholesterol-rich substances under the skin). The effects of cholestasis are severe and widespread, leading to exacerbation of liver disease into systemic disease, liver failure, and the need for liver transplantation. causes of cholestatic liver disease include acute hepatitis and inflammation of the bile ducts.
  • the cholestatic liver disease may include primary biliary cholangitis (PBC), primary sclerosing cholangitis (PSC), progressive familial intrahepatic cholestasis (PFIC), and Alagille syndrome (AS), etc. It is not limited thereto.
  • PBC primary biliary cholangitis
  • PSC primary sclerosing cholangitis
  • PFIC progressive familial intrahepatic cholestasis
  • AS Alagille syndrome
  • Primary biliary cirrhosis also known as primary biliary cholangitis (PBC) is a chronic cholestatic liver disease of unknown etiology. Progressive bile duct damage due to portal and periportal inflammation can lead to progressive fibrosis and eventual cirrhosis of the liver. So far, immunological, genetic and environmental factors are known as potential causes of the disease. Primary biliary cirrhosis is mainly seen in middle-aged women, and the symptoms are fatigue, itchiness, or unexplained hyperlipidemia at the initial onset, and may also appear as symptoms of primary biliary cirrhosis.
  • the treatment method for PBC is bile acid therapy using ursodeoxycholic acid (UDSA) and obeticholic acid (OCA).
  • UDSA ursodeoxycholic acid
  • OCA obeticholic acid
  • the mechanism of action of both drugs in PBC is related to their ability to activate FXR and TGFR-5 to exert anti-inflammatory effects.
  • sufficient biochemical responses were not achieved in about 40% of patients treated with UDCA.
  • PSC Primary sclerosing cholangitis
  • liver function tests using blood show elevated alkaline phosphatase levels, elevated aminotransferase levels, and gamma globulinemia, the diagnosis of primary sclerosing cholangitis is indicated.
  • liver transplantation is the only treatment method that can fundamentally treat it.
  • composition according to the present invention may be characterized in that there is no or a relatively low degree of weight gain, which is a side effect of the existing therapeutic agent for nonalcoholic fatty liver disease.
  • the composition may (a) decrease the expression or activity of collagen-1a, a fibrosis marker; (b) decreased expression or activity of TNF-alpha (tumor necrosis factor- ⁇ ), a pro-inflammatory marker; (c) decreased expression or activity of sterol regulatory element binding protein-1c (SREBP-1c), a lipogenesis marker; (d) reduction of triglycerides in the liver; and (e) one or more of the properties of (a) to (e), such as reducing blood cholesterol, to prevent or treat liver disease.
  • the present invention is not limited thereto.
  • composition can reduce NAS (NAFLD activity score) to prevent or treat liver disease, for example, non-alcoholic fatty liver disease.
  • NAS NAFLD activity score
  • the composition can prevent or treat liver disease, for example, non-alcoholic fatty liver disease by reducing hydroxyprolin, which is known as an indicator of fibrosis in liver tissue.
  • the pharmaceutical composition of the present invention may further include a pharmaceutically acceptable carrier, excipient or diluent.
  • a pharmaceutically acceptable carrier excipient or diluent.
  • Such pharmaceutically acceptable carriers, excipients, or diluents may be non-naturally occurring.
  • the term "pharmaceutically acceptable” means a sufficient amount to exhibit a therapeutic effect and does not cause side effects, and the type of disease, the patient's age, weight, health, sex, and the patient's sensitivity to the drug , administration route, administration method, frequency of administration, treatment period, combination or drugs used at the same time can be easily determined by those skilled in the art according to factors well known in the medical field.
  • the pharmaceutical composition including the peptide of the present invention may further include a pharmaceutically acceptable excipient.
  • the excipient is not particularly limited thereto, but in the case of oral administration, a binder, a lubricant, a disintegrant, a solubilizer, a dispersing agent, a stabilizer, a suspending agent, a dye, a flavoring agent, etc. may be used, and in the case of an injection, a buffer, a preservative, An analgesic agent, a solubilizer, an isotonic agent, a stabilizer, etc. can be mixed and used, and in the case of topical administration, a base, excipient, lubricant, preservative, etc. can be used.
  • the formulation of the composition of the present invention can be prepared in various ways by mixing with the pharmaceutically acceptable excipients as described above.
  • it may be prepared in the form of tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like, and in the case of injections, it may be prepared in the form of unit dose ampoules or multiple doses.
  • it can be formulated as a solution, suspension, tablet, pill, capsule, sustained release formulation, and the like.
  • suitable carriers, excipients and diluents for formulation include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, Microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, or mineral oil may be used.
  • it may further include a filler, an anti-aggregating agent, a lubricant, a wetting agent, a flavoring agent, a preservative, and the like.
  • the pharmaceutical composition of the present invention is any one selected from the group consisting of tablets, pills, powders, granules, capsules, suspensions, internal solutions, emulsions, syrups, sterile aqueous solutions, non-aqueous solvents, freeze-dried preparations and suppositories may have the form of
  • composition is formulated in a dosage form suitable for administration in a patient's body according to a conventional method in the pharmaceutical field, specifically, a formulation useful for administration of a protein drug, and administration commonly used in the art.
  • the triple agonist or its long-acting conjugate can be used by mixing with various pharmaceutically acceptable carriers such as physiological saline or organic solvents, and with glucose, sucrose or dextran to increase stability or absorption.
  • various pharmaceutically acceptable carriers such as physiological saline or organic solvents, and with glucose, sucrose or dextran to increase stability or absorption.
  • Carbohydrates, antioxidants such as ascorbic acid or glutathione, chelating agents, low molecular weight proteins or other stabilizers may be used as agents.
  • the dosage and frequency of administration of the pharmaceutical composition of the present invention is determined according to the type of drug as the active ingredient, together with several related factors such as the disease to be treated, the route of administration, the age, sex and weight of the patient, and the severity of the disease. .
  • the composition of the present invention may include the peptide or a long-acting conjugate comprising the same in a pharmaceutically effective amount, but is not limited thereto.
  • the inclusion of the peptide or the long-acting conjugate in a pharmaceutically effective amount means the degree to which the desired pharmacological activity (eg, prevention, improvement or treatment of liver disease) can be obtained due to the peptide or the long-acting conjugate, and also It may mean a pharmaceutically acceptable level as a level that does not cause toxicity or side effects in the administered subject or is insignificant, but is not limited thereto. Such a pharmaceutically effective amount may be determined by comprehensively considering the number of administration, patient, formulation, and the like.
  • the pharmaceutical composition of the present invention may contain the component (active ingredient) in an amount of 0.01 to 99% by weight to volume.
  • the total effective amount of the composition of the present invention may be administered to a patient as a single dose, or may be administered by a fractionated treatment protocol in which multiple doses are administered for a long period of time.
  • the pharmaceutical composition of the present invention may vary the content of the active ingredient according to the severity of the disease.
  • the preferred total dose of the peptide or conjugate of the present invention may be about 0.0001 mg to 500 mg per 1 kg of the patient's body weight per day, but is not limited thereto.
  • the dosage of the conjugate is determined by considering various factors such as the age, weight, health status, sex, severity of disease, diet and excretion rate of the patient, as well as the administration route and number of treatments of the pharmaceutical composition, and the effective dosage for the patient is determined.
  • the pharmaceutical composition according to the present invention is not particularly limited in its formulation, administration route and administration method as long as the effect of the present invention is exhibited.
  • the triple agonist or long-acting conjugate thereof of the present invention may be an FXR agonist; or an FXR agonist and an ACC inhibitor in an appropriate ratio.
  • the triple agent of the present invention or a long-acting conjugate thereof may be administered in an amount of about 0.1 to about 100 mg, about 0.5 to about 80 mg, about 1 to about 60 mg, about 1 to about 25 mg, administered at about 5 to about 20 mg, or about 10 to about 15 mg
  • the FXR agonist is about 1 to about 100 mg, about 1 to about 250 mg, about 5 to about 250 mg, about 5 to about 50 mg, about 5 to about 40 mg, about 5 to about 40 mg, about 5 to about 25 mg, about 5 to about 30 mg, about 90 to about 250 mg, about 140 to about 250 mg, about 140 to about 200 mg
  • the ACC inhibitor is about 1 to 1000 mg, about 5 to about 1000 mg, about 5 to 150 mg, about 5 to about 50 mg, about 5 to about 30 mg, about 5 to about 750
  • the triple agonist or long-acting conjugate thereof of the present invention may be an FXR agonist; Alternatively, it may be administered in combination with an FXR agonist and an ACC inhibitor in an appropriate ratio to exhibit an excellent therapeutic effect on liver disease.
  • the pharmaceutical composition of the present invention has excellent in vivo persistence and potency, and can significantly reduce the number and frequency of administration of the pharmaceutical preparation of the present invention, but is not limited thereto.
  • Another embodiment embodying the present invention is a substance having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor (triple agonist) or a long-acting conjugate thereof (triple), characterized in that it is used in combination with an FXR agonist It provides a pharmaceutical composition for preventing or treating liver disease, including a long-acting long-acting conjugate).
  • the pharmaceutical composition may be further characterized in that it is used in combination with an ACC inhibitor.
  • a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist, a composition comprising an ACC agonist, prophylaxis, Treatment, liver disease and pharmaceutical compositions are the same as described above.
  • Another aspect embodying the present invention is (i) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), and (ii) administering an FXR agonist to a subject in need thereof.
  • Another aspect embodying the present invention is (i) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) , (ii) an FXR agonist, and (iii) an ACC inhibitor to a subject in need thereof.
  • Another aspect embodying the present invention is a pharmaceutically effective amount of (i) a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonist) or its administering a long-acting conjugate (triple-acting long-acting conjugate); and (ii) administering an FXR agonist, wherein the steps (i) and (ii) are sequential, reversed, or simultaneous.
  • Another aspect embodying the present invention is a pharmaceutically effective amount of (i) a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonist) or its administering a long-acting conjugate (triple-acting long-acting conjugate); (ii) administering an FXR agonist; and (iii) an ACC inhibitor, wherein the steps of (i), (ii), and (iii) are sequential, reversed, or simultaneous.
  • Each step of administering each active ingredient of the prophylactic or therapeutic method of the present invention may be performed in any order, and (i), (ii), and (iii) do not mean the order, and Each step may be performed sequentially, in reverse order, or simultaneously, but is not limited thereto.
  • a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist, a composition comprising an ACC agonist, prophylaxis, Treatment, liver disease and pharmaceutical compositions are the same as described above.
  • the subject is a subject suspected of liver disease
  • the subject suspected of liver disease refers to mammals including mice, livestock, etc. including humans that have or may develop the disease, but the glucagon receptor of the present invention , a substance having activity on the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (a) an FXR agonist, or (b) an FXR agonist and an ACC inhibitor.
  • substances having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor of the present invention triple agonist or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (a) an FXR agonist, or (b) an FXR agonist and an ACC inhibitor, by administering to the subject suspected of liver disease, the subject may be efficiently treated.
  • Liver disease is the same as described above.
  • the method of the present invention comprises a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (a) an FXR agonist, or (b) a combination or pharmaceutical composition comprising an FXR agonist and an ACC inhibitor, in a pharmaceutically effective amount.
  • the method according to the present invention comprises a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (a) the FXR agonist, or (b) the FXR agonist and the ACC inhibitor are administered as one agent, or the separate agents are administered simultaneously, separately, sequentially or in reverse order.
  • the term “administration” means introducing a given substance into a patient (individual) by any suitable method.
  • the route of administration of the triple agonist or long-acting conjugate thereof, FXR agonist, or ACC inhibitor is not particularly limited thereto, but the triple agonist or long-acting conjugate thereof, FXR agonist, or any ACC inhibitor capable of reaching an in vivo target It may be administered through a general route, for example, intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intradermal administration, oral administration, topical administration, intranasal administration, intrapulmonary administration, rectal administration, etc. can be .
  • the triple agonist or long-acting conjugate thereof, FXR agonist, and ACC inhibitor may be administered by the same administration route or may be administered by different administration routes, and the administration routes of the drugs administered in combination are independent of each other can be
  • the triple agonist of the present invention or a long-acting conjugate thereof, an FXR agonist; or a composition comprising a triple agonist or a long-acting combination thereof, an FXR agonist, and an ACC inhibitor once a day, once every 2 days, once every 3 days, once a week, once every two weeks , may be administered once every 4 weeks, or once a month, but is not limited thereto.
  • the FXR agonist and the ACC inhibitor are administered in combination with the triple agonist of the present invention, or a long-acting conjugate thereof, or a composition comprising the same, as long as it can exert a preventive or therapeutic effect on liver disease, but the frequency of administration is not limited, but for example For example, once a day, once every 2 days, once every 3 days, once a week, once every 2 weeks, once every 4 weeks, or once a month.
  • the triple agent or long-acting combination thereof is administered at the same interval, or at a different interval, for example, 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days. It may be administered at intervals of one day, or 10 days or more, but is not limited thereto,
  • a suitable total daily amount may be determined by a treating physician within the scope of sound medical judgment, and may be administered once or divided into several doses.
  • a specific therapeutically effective amount for a particular patient depends on the type and extent of the response to be achieved, the specific composition, including whether other agents are used, if necessary, the specific composition, the patient's age, weight, general health, It is preferable to apply differently depending on various factors including sex and diet, administration time, administration route and secretion rate of the composition, treatment period, drugs used together or concurrently with a specific composition, and similar factors well known in the pharmaceutical field.
  • Another aspect embodying the present invention is a liver disease comprising administering and/or using the combination, a pharmaceutical composition for the prevention or treatment of the disease, or the pharmaceutical kit to an individual in need thereof.
  • a method of prevention or treatment is provided.
  • a combination, kit comprising a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist, and an ACC agonist , composition, prevention, treatment, liver disease and pharmaceutical composition as described above.
  • Another aspect embodying the present invention provides the use of the combination, pharmaceutical composition, or pharmaceutical kit for the prophylaxis or treatment of liver disease and/or the use of a medicament for the prophylaxis or treatment of liver disease. .
  • a combination, kit comprising a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist, and an ACC agonist , composition, prevention, treatment, liver disease and pharmaceutical composition as described above.
  • a triple activator exhibiting activity on all GLP-1 receptors, GIP receptors and glucagon receptors was prepared, and its sequences are shown in Table 1 below.
  • amino acid denoted by X in the sequence shown in Table 1 is Aib (2-aminoisobutyric acid), a non-natural amino acid, and the underlined under the amino acid symbol is the lactam ring between the side chains of the underlined amino acid pair. indicates that it is formed.
  • CA means 4-imidazoacetyl (4-imidazoacetyl).
  • the triple activator of Example 1 (SEQ ID NOs: 21, 22, 42, 43, 50, 77, and 96) for pegylation to the cysteine residues, the molar ratio of the triple activator to maleimide-PEG-aldehyde is 1:1 to 3, and the protein concentration is 1 to 5 mg/ml, and 0.5 to 3 at low temperature. reacted for an hour. At this time, the reaction was carried out in an environment in which 20 to 60% isopropanol was added to 50 mM Tris buffer (pH 7.5).
  • the triple activator was pegylated to the maleimide end of maleimide-PEG-aldehyde by reacting a 10 kDa linearly modified polyethylene glycol, maleimide-PEG-aldehyde (NOF, Japan) with the cysteine residue of the triple activator.
  • reaction solution was applied to SP Sepharose HP (GE healthcare, USA) to purify the tri-activator mono-pegylated to cysteine.
  • the purified mono-pegylated tri-activator and immunoglobulin Fc (homodimer of SEQ ID NO: 138) were mixed in a molar ratio of 1:1 to 5 and a protein concentration of 10 to 50 mg/ml from 4 to The reaction was carried out at 8°C for 12 to 18 hours. The reaction was carried out in an environment in which 10 to 50 mM sodium cyanoborohydride and 10 to 30% isopropanol as reducing agents were added to 100 mM potassium phosphate buffer (pH 6.0).
  • This purified long-acting conjugate has a structure in which a triple active peptide, a polyethylene glycol (PEG) linker and an Fc dimer are covalently linked in a molar ratio of 1:1:1, and the PEG linker is two polypeptide chains of an Fc dimer. It is connected to only one of the chains.
  • PEG polyethylene glycol
  • two monomers having the amino acid sequence of SEQ ID NO: 138 form a homodimer through a disulfide bond between cysteine, which is the 3rd amino acid of each monomer, and the homodimer
  • the monomers of are each independently formed with an internal disulfide bond between cysteines at positions 35 and 95 and an internal disulfide bond between cysteines at positions 141 and 199.
  • the immunoglobulin Fc is an immunoglobulin Fc fragment (a homodimer in which two chains of SEQ ID NO: 138 are linked by a disulfide bond) having a hinge region of the Pro-Ser-Cys-Pro sequence at the N-terminus.
  • International Patent Publication No. WO2007 /021129 was prepared by the method described.
  • conjugate in which the triple activator of SEQ ID NO: 21 and the immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 21 and immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 21'. may be used interchangeably.
  • conjugate in which the triple activator of SEQ ID NO: 22 and immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 22 and immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 22'. may be used interchangeably.
  • conjugate in which the triple activator of SEQ ID NO: 42 and the immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 42 and an immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 42'. may be used interchangeably.
  • conjugate in which the triple activator of SEQ ID NO: 43 and the immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 43 and an immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 43', which may be used interchangeably.
  • conjugate in which the triple activator of SEQ ID NO: 50 and immunoglobulin Fc are linked via PEG was named as 'a conjugate comprising SEQ ID NO: 50 and immunoglobulin Fc' or 'a long-acting conjugate of SEQ ID NO: 50', may be used interchangeably.
  • conjugate in which the triple activator of SEQ ID NO: 77 and the immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 77 and immunoglobulin Fc' or a 'persistent conjugate of SEQ ID NO: 77' may be used interchangeably.
  • conjugate in which the triple activator of SEQ ID NO: 96 and the immunoglobulin Fc are linked through PEG was designated as a 'conjugate comprising SEQ ID NO: 96 and immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 96'. may be used interchangeably.
  • Example 3 Triple activator and long-acting conjugate thereof in vitro activity measurement
  • Each of the above cell lines was transformed to express human GLP-1 receptor, human GCG receptor and human GIP receptor genes in Chinese hamster ovary (CHO), respectively, and is suitable for measuring the activities of GLP-1, GCG and GIP. Therefore, the activity for each part was measured using each transformed cell line.
  • Human GLP-1 was serially diluted from 50 nM to 0.000048 nM by 4 times to measure the GLP-1 activity of the triple activator and its long-acting conjugate prepared in Examples 1 and 2, and prepared in Examples 1 and 2
  • the triple activator and its long-acting conjugate were serially diluted from 400 nM to 0.00038 nM by 4 times.
  • the culture medium was removed from the cultured CHO cells expressing the human GLP-1 receptor, 5 ⁇ l of each serially diluted substance was added to the cells, and then 5 ⁇ l of a buffer containing cAMP antibody was added for 15 minutes. during incubation at room temperature.
  • human GCG was serially diluted from 50 nM to 0.000048 nM by 4 times, and the triple activator prepared in Examples 1 and 2 was serially diluted. and its long-acting conjugate were serially diluted from 400 nM to 0.00038 nM in 4-fold increments.
  • the culture medium was removed from the cultured CHO cells expressing human GCG receptor, 5 ⁇ l of each serially diluted substance was added to the cells, and 5 ⁇ l of a buffer containing cAMP antibody was added thereto, followed by room temperature for 15 minutes.
  • human GIP was serially diluted from 50 nM to 0.000048 nM by 4 times, and the triple activator prepared in Examples 1 and 2 was serially diluted. and its long-acting conjugate were serially diluted from 400 nM to 0.00038 nM in 4-fold increments.
  • the culture medium was removed from the cultured CHO cells expressing the human GIP receptor, 5 ⁇ l of each serially diluted substance was added to the cells, and 5 ⁇ l of a buffer containing cAMP antibody was added thereto, followed by room temperature for 15 minutes.
  • the triple activator prepared above or a long-acting conjugate thereof has a function as a triple activator capable of activating all of the GLP-1 receptor, the GIP receptor and the glucagon receptor, and thus can be used as a therapeutic substance for a desired disease.
  • NASH and fibrosis model a choline deficient, high fat and high cholesterol diet, known as NASH and fibrosis model
  • induced CD-HFD Choline deficient high fat diet
  • the model was induced by performing CD-HFD in C57BL/6 mice for 8 weeks.
  • a long-acting complex of the triple activator of SEQ ID NO: 42 As a representative example of the long-acting complex of the triple activator, a long-acting complex of the triple activator of SEQ ID NO: 42, cilofexor as a representative example of an FXR agonist, and firsocostat as a representative example of an ACC inhibitor Experiments were performed. Kilopexor and pirsocostat were purchased from MCE (MedChemExpress).
  • the induced animal model was an excipient control group, a long-acting conjugate of the triple activator of SEQ ID NO: 42 (2.6 nmo/kg, Q2D, subcutaneous) single administration group, an FXR agonist single administration group (4.3 mmol/kg, QD, oral), ACC inhibitor Single administration group (51.1 mmol/kg, QD, oral), long-acting conjugate of the triple active agent of SEQ ID NO: 42 (2.6 nmo/kg, Q2D, subcutaneous) and FXR agonist (4.3 mmol/kg, QD, oral) combined administration group, Triple active agent of SEQ ID NO: 42 (2.6 nmo/kg, Q2D, subcutaneous) and ACC inhibitor (51.1 mmol/kg, QD, oral) and FXR agonist (4.3 mmol/kg, QD, oral) triple combination administration group , and repeated administration was carried out for 6 weeks. After 6 weeks of repeated administration, H&E staining was performed on the liver tissue of each mouse taken through auto
  • QD means once a day
  • Q2D means once every two days.
  • the NASH therapeutic efficacy of the triple activator or its long-acting conjugate could be further improved by combined administration with an FXR agonist or triple combination administration with an FXR agonist and an ACC inhibitor.
  • the ACC inhibitor and the triple combination administration of the FXR agonist in liver tissue of the CD-HFD mouse model The content of hydroxyproline, known as an indicator of infiltrating fibrosis, was measured.
  • the long-acting complex of the triple activator of SEQ ID NO: 42 which is a representative example of the long-acting conjugate of the triple activator identified above, the group administered alone, the FXR agonist alone group, the ACC inhibitor alone group, the long-acting type of the triple activator of SEQ ID NO: 42
  • the hydroxyproline content was measured in the liver tissue of the CD-HFD mouse model of the group administered with the conjugate and the FXR agonist, and the long-acting conjugate of the triple activator of SEQ ID NO: 42, and the triple combination administration of the ACC inhibitor and FXR agonist.
  • a long-acting conjugate of the triple activator reduced the hydroxyproline content more than the ACC inhibitor and FXR agonist alone group.
  • the hydroxyproline content significantly decreased by the administration of the long-acting conjugate of the triple activator was more effectively reduced during the combined administration with the FXR agonist and the triple combination administration of the ACC inhibitor and the FXR agonist (Fig. 2).
  • the fibrosis therapeutic efficacy of the triple activator of the present invention or its long-acting conjugate could be more effectively improved by combined administration with an FXR agonist or triple combination administration with an FXR agonist and an ACC inhibitor.
  • the triple active agent or long-acting conjugate of the present invention can further improve the therapeutic effect of various nonalcoholic fatty liver diseases by combined administration with an FXR agonist or triple combination administration with an FXR agonist and an ACC inhibitor. This suggests that it can be used as a new combination dosage formulation, or a triple combination dosage formulation.

Abstract

The present invention relates a therapeutic use of a combination containing a farnesoid X receptor agonist and a triple agonistic long-acting conjugate or a triple agonist, or a combination further containing an acetyl-CoA carboxylase inhibitor in addition to the above.

Description

3중 작용성 지속형 결합체 또는 3중 작용제를 포함하는 조합물의 치료학적 용도Therapeutic use of triple-acting long-acting conjugates or combinations comprising triple agents
본 발명은 3중 작용성 지속형 결합체 (triple agonistic long-acting conjugate) 또는 3중 작용제 (triple agonist); 및 FXR 작용제 (farnesoid X receptor agonist)를 포함하는 조합물, 또는 상기 조합물에 ACC 억제제 (acetyla-CoA carboxylase inhibitor)를 추가로 포함하는 조합물의 치료학적 용도에 관한 것이다.The present invention provides a triple agonistic long-acting conjugate or a triple agonist; and an FXR agonist (farnesoid X receptor agonist), or to the therapeutic use of the combination further comprising an acetyla-CoA carboxylase inhibitor (ACC) in the combination.
간은 동물의 주요 생체 기관 중 하나로, 간과 관련된 질환으로는 대표적으로 비알코올성 지방간, 간염, 간섬유증, 담즙정체성 간질환, 간경변, 간 부전, 간암, 등을 들 수 있다. 바이러스, 알코올, 약물, 면역 이상, 대사 질환 등의 원인으로 인해 간에 염증이 발생할 수 있고, 간염증의 진행 및 만성화에 따라 간섬유증, 간경변, 간암 간암 등의 질환으로 발병하는 것으로 알려져 있다.The liver is one of the main living organs of animals, and liver-related diseases include nonalcoholic fatty liver, hepatitis, liver fibrosis, cholestatic liver disease, cirrhosis, liver failure, liver cancer, and the like. Inflammation of the liver may occur due to causes such as viruses, alcohol, drugs, immune abnormalities, metabolic diseases, and the like, and it is known that diseases such as liver fibrosis, cirrhosis, liver cancer and liver cancer occur as hepatitis progresses and becomes chronic.
일반적으로 간에 염증이 생기는 간염이 간 질환의 대부분을 차지하며, 간염증이 진행되면서 간 염증을 수반하거나, 또는 간 염증에 기인하여 다양한 간질환(간섬유증, 경변 등)이 나타나는 것으로 알려져 있다. 간염증의 양상에 따라 급성 간염과 만성 간염, 원인에 따라 바이러스성 간염, 알코올성 간염, 약물성 간염 등으로 나눌 수 있다. 담즙정체성 간질환(cholestasis liver disease) 역시 염증성 질환이 원인이 되어, 발생하는 것으로 추정되고 있다.In general, hepatitis, which is caused by inflammation of the liver, accounts for most of the liver diseases, and it is known that various liver diseases (liver fibrosis, cirrhosis, etc.) appear due to liver inflammation or liver inflammation as hepatitis progresses. According to the pattern of hepatitis, acute hepatitis and chronic hepatitis, depending on the cause, can be divided into viral hepatitis, alcoholic hepatitis, drug hepatitis, and the like. Cholestatic liver disease (cholestasis liver disease) is also assumed to be caused by an inflammatory disease.
한편, 비알코올성 지방간 질환 (NAFLD, nonalcoholic fatty liver disease)은 알코올 섭취와 관련 없음에도 알코올 간염과 유사한 조직소견을 보이는 질환의 일종으로, 단순 지방증(simple steatosis), 비알코올성 지방간 (non-alcoholic fatty liver, NAFL)으로부터 비알코올성지방간염 (non-acoholic steatohepatitis, NASH), 간 염증, 간 섬유증 (liver fibrosis), 간경변 (cirrhosis), 간부전 (liver decompensation), 및 간암 (hepatocellular carcinomas)까지를 아우르는 질환이다. 비알코올성 지방간 질환은 비만 및 당뇨인구가 증가함에 따라 함께 증가하고 있는 추세이다.On the other hand, nonalcoholic fatty liver disease (NAFLD) is a type of disease that shows tissue findings similar to alcoholic hepatitis even though it is not related to alcohol intake. Simple steatosis, non-alcoholic fatty liver , NAFL) to non-acoholic steatohepatitis (NASH), liver inflammation, liver fibrosis, cirrhosis, liver decompensation, and hepatocellular carcinomas. Nonalcoholic fatty liver disease is on the rise as the obesity and diabetic populations increase.
이러한 비알코올성 지방간질환의 병인으로는 인슐린 저항성, 비만, 지질독성 (lipotoxicity), 염증 반응등과 같은 다양한 병인에 기인하다고 알려져 있다. 그 중 가장 큰 병인으로는 인슐린 저항성을 꼽는다.It is known that the etiology of such nonalcoholic fatty liver disease is due to various etiologies such as insulin resistance, obesity, lipotoxicity, and inflammatory response. One of the most important causes is insulin resistance.
이와 같은 비알코올성 지방간질환을 예방 및/또는 치료하기 위해 인슐린 저항성을 개선시키는데 많은 노력을 하고 있다. 예를 들어, 인슐린 반응개선제 (insulin sensitizer)의 일종인 TZD (thiazolidnedinones)나 메포민에 대한 임상시험은 현재에도 활발하게 이루어지고 있다 (Hepatology(2003) 38:1008-17, J Clin Invest. (2001) 108:1167-74).In order to prevent and/or treat such nonalcoholic fatty liver disease, much effort is being made to improve insulin resistance. For example, clinical trials for thiazolidnedinones (TZD) or metformin, a type of insulin sensitizer, are still being actively conducted (Hepatology (2003) 38: 1008-17, J Clin Invest. (2001) ) 108:1167-74).
그러나 TZD계열의 약물을 이용한 치료의 경우 체중증가가 크고, 체액 흐름을 느리게 할 수 있다는 단점이 있어 심장질환 환자에게는 적용이 불가능한 것으로 알려져 있다. TZD 계열의 약물 외에도 빅토자 (Victoza) 또는 바이에타 (Byetta)와 같은 GLP-1 수용체 아고니스트를 이용한 비알코올성 지방간질환에 대한 적용 임상실험도 활발히 이루어지고 있다. 특히, 해당약물은 체중감량 효과를 수반하기 때문에 개발초기에는 비알코올성 지방간질환 및 이에 기인한 섬유증에도 치료효과가 있을 것으로 예상되었다. 하지만, 이들의 경우는 다른 펩타이드 호르몬과 마찬가지로 체내 반감기가 극히 짧아 매일 또는 하루 두번 이상의 반복투여를 해야하기 때문에, 환자 편의성에 대한 단점이 있다. 이와 같은 잦은 투여는 환자에게 엄청난 고통과 불편함을 야기하게 된다. 최근에는 이러한 투여편의성이 개선된 GLP-1 수용체 아고니스트인 세마글루타이드 (semaglutide)에 대한 비알코올성 지방간질환 임상실험이 진행되었으나, 여전히 비알코올성 지방간염에서 기인한 섬유증에 대한 치료효능이 없음이 확인되었다. 즉, 단순히 인슐린 저항성을 개선시키는 기작 또는 GLP-1 수용체 아고니스트를 포함하는 다양한 당뇨병 치료제의 비알코올성 지방간질환 치료제로의 적용은 여전히 부족한 치료효능, 다양한 부작용, 그리고 환자 편의성 문제 등의 단점들이 있다. 이러한 결과 등으로 인하여, 당뇨병 치료에 효과가 있다고 알려진 약물을 비알코올성 지방간질환의 치료제로 직접 사용하기는 어렵다는 사실이 당업계에는 다양하게 알려져 있다. 따라서, 당뇨병 치료에 효과가 있다고 알려진 약물이 곧 비알코올성 지방간질환의 치료제로 사용될 수 있는지 여부는 임상시험을 통한 상기 질환의 치료효능을 명확히 확인한 후에야 결정할 수 있을 것이다. 하지만, 현재 사용중인 당뇨병치료제 중 비알코올성 지방간질환 치료제로 승인을 받은 약물은 전무한 실정이다. 따라서, 환자 편의성 확보 및 부작용 없이 보다 효과적으로 비알코올성 지방간질환을 치료할 수 있는 약물의 개발이 여전히 요구되고 있다.However, it is known that treatment using TZD-type drugs is not applicable to patients with heart disease because of the disadvantages of large weight gain and slowing the flow of body fluids. In addition to TZD drugs, clinical trials for nonalcoholic fatty liver disease using GLP-1 receptor agonists such as Victoza or Byetta are also being actively conducted. In particular, the drug was expected to have a therapeutic effect on nonalcoholic fatty liver disease and fibrosis caused by it in the early stage of development because it is accompanied by a weight loss effect. However, in these cases, like other peptide hormones, the half-life in the body is extremely short, and since repeated administration is required every day or twice a day, there is a disadvantage in patient convenience. Such frequent administration causes great pain and discomfort to the patient. Recently, clinical trials for nonalcoholic fatty liver disease have been conducted on semaglutide, a GLP-1 receptor agonist with improved administration convenience, but it is still confirmed that there is no therapeutic effect on fibrosis caused by nonalcoholic steatohepatitis. became In other words, the application of various antidiabetic drugs including a mechanism to simply improve insulin resistance or a GLP-1 receptor agonist to a therapeutic agent for nonalcoholic fatty liver disease still lacks therapeutic efficacy, various side effects, and In addition, there are disadvantages such as the problem of patient convenience. Due to these results and the like, it is variously known in the art that it is difficult to directly use a drug known to be effective in treating diabetes as a therapeutic agent for nonalcoholic fatty liver disease. Therefore, whether a drug known to be effective in treating diabetes can be used as a therapeutic agent for nonalcoholic fatty liver disease will be determined only after clearly confirming the therapeutic efficacy of the disease through clinical trials. However, none of the currently used diabetes treatment drugs have been approved as a treatment for nonalcoholic fatty liver disease. Therefore, there is still a need to develop a drug that can more effectively treat non-alcoholic fatty liver disease without securing patient convenience and side effects.
GLP-1 (Glucagon-like peptide-1) 및 GIP (Glucose-dependent insuliontropic polypeptide)는 대표적인 위장 호르몬이자 신경 호르몬으로서 음식물 섭취에 따른 혈중 당성분 조절에 관여하는 물질이다. 글루카곤 (Glucagon)은 췌장에서 분비되는 펩타이드 호르몬으로 전술한 두 물질과 함께 혈중 당 농도 조절 작용에 관여한다.GLP-1 (Glucagon-like peptide-1) and GIP (Glucose-dependent insuliontropic polypeptide) are representative gastrointestinal hormones and neurohormones that are involved in the regulation of blood sugar components according to food intake. Glucagon is a peptide hormone secreted by the pancreas and is involved in blood sugar concentration control together with the two substances described above.
한편, GLP-1는 비만 치료제로서의 가능성을, GIP는 혈당 농도에 의존적으로 췌장에서의 인슐린 분비를 촉진하고 혈당 농도를 낮추는데 도움을 주는 기능을 수행하며 GLP-1의 활성 증가 효과, 항염증 효과등이 보고되었다. 글루카곤은 약물 치료 또는 질병, 호르몬이나 효소 결핍 등의 원인으로 혈당이 떨어지기 시작하면 췌장에서 생산된다. 글루카곤은 간에 신호하여 글리코겐을 분해하여 글루코오스를 방출하도록 유도하고, 혈당 수준을 정상 수준까지 높이는 역할을 한다. 뿐만 아니라, 글루카곤은 혈당 상승효과 이외에 동물과 사람에서의 식욕억제 및 지방세포의 호르몬 민감성 리파제 (hormone sensitive lipase)를 활성화시켜 지방 분해를 촉진 및 에너지대사 (energy expenditure)를 촉진하여 항비만 효과를 나타냄이 보고되었다. On the other hand, GLP-1 has potential as a therapeutic agent for obesity, and GIP functions to promote insulin secretion from the pancreas and to lower blood sugar concentration dependent on blood sugar concentration, and increases the activity of GLP-1, anti-inflammatory effects, etc. This has been reported Glucagon is produced by the pancreas when blood sugar begins to drop due to a cause, such as medication or disease, or a hormone or enzyme deficiency. Glucagon signals the liver to break down glycogen to release glucose, and plays a role in raising blood sugar levels to normal levels. In addition, glucagon exhibits anti-obesity effects by promoting lipolysis and energy expenditure by activating hormone-sensitive lipase in adipocytes and suppressing appetite in animals and humans, in addition to raising blood sugar. This has been reported
FXR 작용제 (farnesoid X receptor agonist)는 담즙산 수용체(Bile acid Receptor, BAR)로도 공지되어 있는, 담즙산에 의해 활성화되는 핵 수용체인 FXR 작용제이다. FXR은 간, 장 및 신장과 같은 담즙산 대사의 주요 부위에서 발현되며, 이는 조직-특이적 방식으로 작용하여 담즙산을 포함한 여러 대사 경로는 물론, 섬유화 작용에도 영향을 미친다고 보고되어있다. 이에 FXR 에 의해 매개되는 간 질환에 대한 효과적인 치료 방법이 요구되고 있다.FXR agonists (farnesoid X receptor agonists) are FXR agonists, which are nuclear receptors activated by bile acids, also known as bile acid receptors (BARs). FXR is expressed in major sites of bile acid metabolism such as liver, intestine and kidney, and it has been reported that it acts in a tissue-specific manner to affect several metabolic pathways including bile acids as well as fibrosis. Accordingly, there is a need for an effective treatment method for liver disease mediated by FXR.
ACC 억제제 (acetyla-CoA carboxylase inhibitor)는 지방산 생성 및 대사의 조절에 중요한 효소인 아세틸-CoA 카복실라제 (acetyla-CoA carboxylase)를 억제하는 물질로 알려져 있다.ACC inhibitors (acetyla-CoA carboxylase inhibitors) are known as substances that inhibit acetyla-CoA carboxylase, an enzyme important in the regulation of fatty acid production and metabolism.
효과적인 간질환의 예방 및/또는 치료를 위한 약물의 개발이 요구된다.The development of drugs for the effective prevention and/or treatment of liver disease is required.
본 발명의 하나의 목적은 약학적 유효량의 3중 작용성 지속형 결합체(triple agonistic long-acting conjugate) 또는 3중 작용제 (triple agonist)와 약학적으로 허용되는 부형제를 함유하는 간 질환의 치료 또는 예방을 위한 약학적 조성물로서, 상기 약학적 조성물은 파네소이드 X 수용체 작용제(farnesoid X receptor agonist)와 병용되는 것을 특징으로 하는 약학적 조성물을 제공하는 것이다.One object of the present invention is to treat or prevent liver disease containing a pharmaceutically effective amount of a triple agonistic long-acting conjugate or a triple agonist and a pharmaceutically acceptable excipient. As a pharmaceutical composition for
본 발명의 또 하나의 목적은 (i) 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 갖는 물질 또는 이의 지속형 결합체, 및 (ii) 파네소이드 X 수용체 작용제 (farnesoid X receptor agonist, FXR 작용제)를 포함하는, 조합물을 제공하는 것이다.Another object of the present invention is (i) a substance having activity on glucagon receptor, GLP-1 (Glucagon-like peptide-1) receptor, and GIP (glucose-dependent insuliontropic polypeptide) receptor, or a long-acting conjugate thereof, and (ii) a farnesoid X receptor agonist (FXR agonist).
본 발명의 또 하나의 목적은 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 또는 이의 지속형 결합체, (ii) FXR 작용제, 및 (iii) 아세틸조효소 A 카르복실라제 억제제 (acetyla-CoA carboxylase inhibitor, ACC 억제제)를 포함하는, 조합물을 제공하는 것이다.Another object of the present invention is (i) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor or a long-acting conjugate thereof, (ii) an FXR agonist, and (iii) an acetyl coenzyme A carboxylase To provide a combination comprising an inhibitor (acetyla-CoA carboxylase inhibitor, ACC inhibitor).
본 발명의 또 하나의 목적은 상기 조합물을 포함하는, 간 질환의 예방 또는 치료용 약학적 조성물을 제공하는 것이다.Another object of the present invention is to provide a pharmaceutical composition for preventing or treating liver disease, comprising the combination.
본 발명의 또 하나의 목적은 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 또는 이의 지속형 결합체, 및 (ii) FXR 작용제 (farnesoid X receptor agonist)를 포함하는, 간 질환의 예방 또는 치료를 위한 약학적 키트를 제공하는 것이다.Another object of the present invention comprises (i) a substance having activity on glucagon receptor, GLP-1 receptor, and GIP receptor or a long-acting conjugate thereof, and (ii) FXR agonist (farnesoid X receptor agonist), To provide a pharmaceutical kit for the prevention or treatment of liver disease.
본 발명의 또 하나의 목적은 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 또는 이의 지속형 결합체, (ii) FXR 작용제, 및 (iii) ACC 억제제를 포함하는, 간 질환의 예방 또는 치료를 위한 약학적 키트를 제공하는 것이다.Another object of the present invention is comprising (i) a substance having activity on glucagon receptor, GLP-1 receptor, and GIP receptor or a long-acting conjugate thereof, (ii) an FXR agonist, and (iii) an ACC inhibitor, To provide a pharmaceutical kit for the prevention or treatment of liver disease.
본 발명의 또 하나의 목적은 상기 조합물, 상기 간 질환의 치료 또는 예방을 위한 약학적 조성물, 또는 상기 약학적 키트를 이를 필요로 하는 개체에 투여 및/또는 사용하는 단계를 포함하는 간 질환의 예방 또는 치료 방법을 제공하는 것이다.Another object of the present invention is to treat liver disease comprising administering and/or using the combination, a pharmaceutical composition for the treatment or prevention of liver disease, or the pharmaceutical kit to an individual in need thereof. To provide a method of prevention or treatment.
본 발명의 또 하나의 목적은 상기 조합물, 약학적 조성물, 또는 약학적 키트의 간 질환의 예방 또는 치료 용도 및/또는 비알코올성 지방간 질환의 예방 또는 치료를 위한 약제의 제조를 위한 용도를 제공하는 것이다.Another object of the present invention is to provide use of the combination, pharmaceutical composition, or pharmaceutical kit for the prophylaxis or treatment of liver disease and/or use for the preparation of a medicament for the prevention or treatment of non-alcoholic fatty liver disease. will be.
본 발명에 따른 3중 작용성 지속형 결합체 또는 3중 작용제; 및 FXR 작용제의 복합 병용 투여, 또는 3중 작용성 지속형 결합체 또는 3중 작용제; FXR 작용제; 및 ACC 억제제의 삼중 복합 투여 요법은 단일 투여 요법에 비해 향상된 효과를 가져서 간 질환의 예방 또는 치료에 유용하게 사용할 수 있다.The triple acting long-acting conjugate or triple agent according to the present invention; and concomitant administration of an FXR agonist, or a triple acting long-acting conjugate or a triple agent; FXR agonists; And the triple combination administration regimen of an ACC inhibitor has an improved effect compared to a single administration regimen, so it can be usefully used for the prevention or treatment of liver disease.
도 1은 삼중 활성체의 지속형 결합체의 복합 투여에 따른 NAS (NAFLD activity score)의 변화를 나타낸 도이다 (*p < 0.05, **p < 0.01, ***p < 0.001 vs. CH-HFD, 부형제 대조군 by ono-way ANOVA).1 is a diagram showing the change in NAS (NAFLD activity score) according to the complex administration of the long-acting conjugate of the triple activator (*p < 0.05, **p < 0.01, ***p < 0.001 vs. CH-HFD , vehicle control by ono-way ANOVA).
도 2는 삼중 활성체의 지속형 결합체의 복합 투여에 따른 간조직내 하이드록시프롤린 (hydroxyprolin)의 함량 변화를 나타낸 도이다 (*p < 0.05, **p < 0.01, ***p < 0.001 vs. CH-HFD, 부형제 대조군 by ono-way ANOVA).2 is a diagram showing the change in the content of hydroxyprolin in liver tissue according to the complex administration of the long-acting conjugate of the triple activator (*p < 0.05, **p < 0.01, ***p < 0.001 vs. CH-HFD, vehicle control by ono-way ANOVA).
본 발명을 구현하는 하나의 양태는 3중 작용성 지속형 결합체(triple agonistic long-acting conjugate) 또는 3중 작용제 (triple agonist)를 포함하는 간 질환의 치료 또는 예방을 위한 약학적 조성물로서, 상기 약학적 조성물은 파네소이드 X 수용체 작용제(farnesoid X receptor agonist)와 병용되는 것을 특징하는 조성물이다.One aspect embodying the present invention is a pharmaceutical composition for the treatment or prevention of liver disease comprising a triple agonistic long-acting conjugate or a triple agonist, the pharmaceutical composition The red composition is a composition characterized in that it is used in combination with a farnesoid X receptor agonist.
하나의 구체예로서, 상기 약학적 조성물은 약학적 유효량의 3중 작용성 지속형 결합체 또는 3중 작용제 (triple agonist)와 약학적으로 허용되는 부형제를 함유하는 간 질환의 치료 또는 예방을 위한 약학적 조성물로서,In one embodiment, the pharmaceutical composition contains a pharmaceutically effective amount of a triple-acting long-acting conjugate or a triple agonist and a pharmaceutically acceptable excipient for the treatment or prevention of liver disease. As a composition,
상기 약학적 조성물은 파네소이드 X 수용체 작용제(farnesoid X receptor agonist)와 병용되는 것을 특징으로 하고,The pharmaceutical composition is characterized in that it is used in combination with a farnesoid X receptor agonist,
상기 3중 작용성 지속형 결합체는 하기 화학식 1로 표시되는 물질인 약학적 조성물인 것을 특징으로 한다: The triple-acting long-acting conjugate is characterized in that the pharmaceutical composition is a substance represented by the following formula (1):
[화학식 1][Formula 1]
Z - Lx - FcZ - Lx - Fc
이 때 Lx는 에틸렌글리콜 반복 단위를 함유하는 링커이며, x는 0 또는 자연수이고,In this case, Lx is a linker containing an ethylene glycol repeating unit, and x is 0 or a natural number,
Fc는 면역글로불린 Fc 영역이고, Fc is an immunoglobulin Fc region,
-는 Lx와 Z 사이, Fc와 Lx 사이의 각각에 공유결합 연결을 나타내며,- represents a covalent linkage between Lx and Z, between Fc and Lx, respectively,
Z 또는 3중 작용제는 하기 일반식 1로 표시되는 아미노산 서열을 포함하는 펩타이드이고, Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 1,
Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1(일반식 1, 서열번호 103)Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp23 Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (Formula 1, SEQ ID NO: 103)
상기 일반식 1에서,In the above general formula 1,
Xaa1은 히스티딘(His, H), 4-이미다조아세틸(CA) 또는 티로신(Tyr, Y)이고, Xaa1 is histidine (His, H), 4-imidazoacetyl (CA) or tyrosine (Tyr, Y),
Xaa2는 글리신(Gly, G), 알파-메틸-글루탐산 또는 2-아미노이소부티르산(2-aminoisobutyric acid, Aib)이며,Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid or 2-aminoisobutyric acid (Aib),
Xaa3은 글루탐산(Glu, E) 또는 글루타민(Gln, Q)이고, Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q),
Xaa7은 트레오닌(Thr, T) 또는 이소류신(Ile, I)이며,Xaa7 is threonine (Thr, T) or isoleucine (Ile, I),
Xaa10은 류신(Leu, L), 티로신(Tyr, Y), 리신(Lys, K), 시스테인(Cys, C) 또는 발린(Val, V)이고, Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C) or valine (Val, V);
Xaa12는 리신(Lys, K), 세린(Ser, S) 또는 이소류신(Ile, I)이며, Xaa12 is lysine (Lys, K), serine (Ser, S) or isoleucine (Ile, I);
Xaa13은 글루타민(Gln, Q), 티로신(Tyr, Y), 알라닌(Ala, A) 또는 시스테인(Cys, C)이고, Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A) or cysteine (Cys, C);
Xaa14는 류신(Leu, L), 메티오닌(Met, M) 또는 티로신(Tyr, Y)이며,Xaa14 is leucine (Leu, L), methionine (Met, M) or tyrosine (Tyr, Y);
Xaa15는 시스테인(Cys, C), 아스파르트산(Asp, D), 글루탐산(Glu, E) 또는 류신(Leu, L)이며,Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E) or leucine (Leu, L),
Xaa16은 글리신(Gly, G), 글루탐산(Glu, E) 또는 세린(Ser, S)이고, Xaa16 is glycine (Gly, G), glutamic acid (Glu, E) or serine (Ser, S);
Xaa17은 글루타민(Gln, Q), 아르기닌(Arg, R), 이소류신(Ile, I), 글루탐산(Glu, E), 시스테인(Cys, C) 또는 리신(Lys, K)이며,Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C) or lysine (Lys, K);
Xaa18은 알라닌(Ala, A), 글루타민(Gln, Q), 아르기닌(Arg, R) 또는 히스티딘(His, H)이고,Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R) or histidine (His, H);
Xaa19는 알라닌(Ala, A), 글루타민(Gln, Q), 시스테인(Cys, C) 또는 발린(Val, V)이며,Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C) or valine (Val, V);
Xaa20은 리신(Lys, K), 글루타민(Gln, Q) 또는 아르기닌(Arg, R)이고,Xaa20 is lysine (Lys, K), glutamine (Gln, Q) or arginine (Arg, R);
Xaa21은 글루탐산(Glu, E), 글루타민(Gln, Q), 류신(Leu, L), 시스테인(Cys, C) 또는 아스파르트산(Asp, D)이며,Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C) or aspartic acid (Asp, D);
Xaa23은 이소류신(Ile, I) 또는 발린(Val, V)이고,Xaa23 is isoleucine (Ile, I) or valine (Val, V),
Xaa24는 알라닌(Ala, A), 글루타민(Gln, Q), 시스테인(Cys, C), 아스파라긴(Asn, N), 아스파르트산(Asp, D) 또는 글루탐산(Glu, E)이며,Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D) or glutamic acid (Glu, E),
Xaa27은 발린(Val, V), 류신(Leu, L) 또는 리신(Lys, K) 이고,Xaa27 is valine (Val, V), leucine (Leu, L) or lysine (Lys, K);
Xaa28은 시스테인(Cys, C), 리신(Lys, K), 알라닌(Ala, A), 아스파라긴(Asn, N) 또는 아스파르트산(Asp, D)이며,Xaa28 is cysteine (Cys, C), lysine (Lys, K), alanine (Ala, A), asparagine (Asn, N) or aspartic acid (Asp, D);
Xaa29는 시스테인(Cys, C), 글리신(Gly, G), 글루타민(Gln, Q), 트레오닌(Thr, T), 글루탐산(Glu, E) 또는 히스티딘(His, H)이고,Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q), threonine (Thr, T), glutamic acid (Glu, E) or histidine (His, H);
Xaa30은 시스테인(Cys, C), 글리신(Gly, G), 리신(Lys, K) 또는 히스티딘(His, H)이거나, 부존재하며,Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K) or histidine (His, H), or is absent;
R1은 시스테인(Cys, C), GKKNDWKHNIT(서열번호 106), m-SSGAPPPS-n(서열번호 107) 또는 m-SSGQPPPS-n(서열번호 108)이거나, 부존재하며, R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107) or m-SSGQPPPS-n (SEQ ID NO: 108), or is absent;
m은 -Cys-, -Pro- 또는 -Gly-Pro-이고, m is -Cys-, -Pro- or -Gly-Pro-;
n은 부존재하거나 -Cys-, -Gly-, -Ser- 또는 -His-Gly-이다.n is absent or is -Cys-, -Gly-, -Ser- or -His-Gly-.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 일반식 1에서As a pharmaceutical composition according to any one of the preceding embodiments, in the general formula 1
Xaa2는 글리신, 알파-메틸-글루탐산 또는 2-아미노이소부티르산이고,Xaa2 is glycine, alpha-methyl-glutamic acid or 2-aminoisobutyric acid,
Xaa7은 트레오닌이며,Xaa7 is threonine,
Xaa10은 티로신, 시스테인 또는 발린이고, Xaa10 is tyrosine, cysteine or valine,
Xaa12는 리신 또는 이소류신이며, Xaa12 is lysine or isoleucine,
Xaa13은 티로신, 알라닌, 글루타민 또는 시스테인이고, Xaa13 is tyrosine, alanine, glutamine or cysteine,
Xaa14는 류신, 티로신 또는 메티오닌이며;Xaa14 is leucine, tyrosine or methionine;
Xaa15는 시스테인, 류신, 글루탐산 또는 아스파르트산이고,Xaa15 is cysteine, leucine, glutamic acid or aspartic acid,
Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 글루탐산 또는 리신이며,Xaa17 is glutamine, arginine, isoleucine, cysteine, glutamic acid or lysine,
Xaa18은 알라닌, 글루타민, 아르기닌 또는 히스티딘이고,Xaa18 is alanine, glutamine, arginine or histidine;
Xaa19는 알라닌, 글루타민, 발린 또는 시스테인이며,Xaa19 is alanine, glutamine, valine or cysteine,
Xaa20은 리신, 아르기닌 또는 글루타민이고,Xaa20 is lysine, arginine or glutamine,
Xaa21은 글루탐산, 글루타민, 류신, 시스테인 또는 아스파르트산이며,Xaa21 is glutamic acid, glutamine, leucine, cysteine or aspartic acid,
Xaa23은 이소류신 또는 발린이고,Xaa23 is isoleucine or valine,
Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 글루탐산 또는 아스파르트산이며,Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid or aspartic acid;
Xaa27은 류신 또는 리신인 것을 특징으로 한다.Xaa27 is characterized in that it is leucine or lysine.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 Z 또는 3중 작용제는 하기 일반식 2로 표시되는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다:The pharmaceutical composition according to any one of the preceding embodiments, wherein the Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 2:
Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31- Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40(일반식 2, 서열번호 104)Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (Formula 2, SEQ ID NO: 104)
상기 일반식 2에서,In the above general formula 2,
Xaa1은 4-이미다조아세틸, 히스티딘 또는 티로신이고,Xaa1 is 4-imidazoacetyl, histidine or tyrosine;
Xaa2는 글리신, 알파-메틸-글루탐산 또는 2-아미노이소부티르산이며,Xaa2 is glycine, alpha-methyl-glutamic acid or 2-aminoisobutyric acid;
Xaa10은 티로신 또는 시스테인이고,Xaa10 is tyrosine or cysteine,
Xaa13은 알라닌, 글루타민, 티로신 또는 시스테인이며,Xaa13 is alanine, glutamine, tyrosine or cysteine,
Xaa14는 류신, 메티오닌 또는 티로신이고,Xaa14 is leucine, methionine or tyrosine,
Xaa15는 아스파르트산, 글루탐산 또는 류신이며,Xaa15 is aspartic acid, glutamic acid or leucine,
Xaa16은 글리신, 글루탐산 또는 세린이고,Xaa16 is glycine, glutamic acid or serine,
Xaa17은 글루타민, 아르기닌, 이소류신, 글루탐산, 시스테인 또는 리신이며,Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine or lysine,
Xaa18은 알라닌, 글루타민, 아르기닌 또는 히스티딘이고,Xaa18 is alanine, glutamine, arginine or histidine;
Xaa19는 알라닌, 글루타민, 시스테인 또는 발린이며,Xaa19 is alanine, glutamine, cysteine or valine,
Xaa20은 리신, 글루타민 또는 아르기닌이고,Xaa20 is lysine, glutamine or arginine,
Xaa21은 시스테인, 글루탐산, 글루타민, 류신 또는 아스파르트산이며,Xaa21 is cysteine, glutamic acid, glutamine, leucine or aspartic acid,
Xaa23은 이소류신 또는 발린이고,Xaa23 is isoleucine or valine,
Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴 또는 글루탐산이며,Xaa24 is cysteine, alanine, glutamine, asparagine or glutamic acid,
Xaa28은 리신, 시스테인, 아스파라긴 또는 아스파르트산이고,Xaa28 is lysine, cysteine, asparagine or aspartic acid;
Xaa29는 글리신, 글루타민, 시스테인 또는 히스티딘이며,Xaa29 is glycine, glutamine, cysteine or histidine;
Xaa30은 시스테인, 글리신, 리신 또는 히스티딘이고,Xaa30 is cysteine, glycine, lysine or histidine;
Xaa31은 프롤린 또는 시스테인이며;Xaa31 is proline or cysteine;
Xaa40은 시스테인이거나, 부존재한다.Xaa40 is cysteine or absent.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 Z 또는 3중 작용제는 하기 일반식 3으로 표시되는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다:The pharmaceutical composition according to any one of the preceding embodiments, wherein the Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 3:
Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40(일반식 3, 서열번호 105),Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp- Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (general formula 3, SEQ ID NO: 105);
상기 일반식 3에서,In the above general formula 3,
Xaa1은 히스티딘 또는 티로신이고,Xaa1 is histidine or tyrosine,
Xaa2는 알파-메틸-글루탐산 또는 2-아미노이소부티르산이며,Xaa2 is alpha-methyl-glutamic acid or 2-aminoisobutyric acid,
Xaa13은 알라닌, 티로신 또는 시스테인이고,Xaa13 is alanine, tyrosine or cysteine,
Xaa17은 아르기닌, 시스테인 또는 리신이며,Xaa17 is arginine, cysteine or lysine,
Xaa18은 알라닌 또는 아르기닌이고,Xaa18 is alanine or arginine,
Xaa19는 알라닌 또는 시스테인이며,Xaa19 is alanine or cysteine,
Xaa21은 글루탐산 또는 아스파르트산이고,Xaa21 is glutamic acid or aspartic acid,
Xaa24는 글루타민 또는 아스파라긴이며,Xaa24 is glutamine or asparagine,
Xaa28은 시스테인 또는 아스파르트산이고,Xaa28 is cysteine or aspartic acid,
Xaa29는 시스테인, 히스티딘 또는 글루타민이며,Xaa29 is cysteine, histidine or glutamine;
Xaa30은 시스테인 또는 히스티딘이고,Xaa30 is cysteine or histidine,
Xaa31은 프롤린 또는 시스테인이며,Xaa31 is proline or cysteine,
Xaa40은 시스테인 또는 부존재함.Xaa40 is cysteine or absent.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 Z 또는 3중 작용제 중 상기 Xaa16이 글루탐산이고, Xaa20이 리신이며, 상기 글루탐산과 상기 리신이 락탐 고리를 형성한 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein among the Z or triple agents, Xaa16 is glutamic acid, Xaa20 is lysine, and the glutamic acid and the lysine form a lactam ring.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 Z 또는 3중 작용제는 그 C-말단이 아미드화된 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the Z or triple agent is amidated at its C-terminus.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 Lx 내의 에틸렌글리콜 반복 단위 부분의 화학식량은 1 ~ 100 kDa 범위 내인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the formula weight of the ethylene glycol repeating unit moiety in Lx is in the range of 1 to 100 kDa.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 Z 또는 3중 작용제는 서열번호: 1 내지 102로 이루어진 군으로부터 선택하는 하나의 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the Z or triple agent is a peptide comprising one amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 Z 또는 3중 작용제는 서열번호 21, 22, 42, 43, 50, 64, 66, 67, 70, 71, 76, 77, 96, 97과 100으로 이루어진 군으로부터 선택하는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the Z or triple agent is SEQ ID NO: 21, 22, 42, 43, 50, 64, 66, 67, 70, 71, 76, 77, 96, 97 and It is characterized in that it is a peptide comprising an amino acid sequence selected from the group consisting of 100.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 Z 또는 3중 작용제는 서열번호 21, 22, 42, 43, 50, 66, 67, 77, 96, 97과 100으로 이루어진 군으로부터 선택하는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the Z or triple agent is an amino acid selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 66, 67, 77, 96, 97 and 100 It is characterized in that it is a peptide comprising a sequence.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 Z 또는 3중 작용제는 서열번호 21, 22, 42, 43, 50, 77과 96으로 이루어진 군으로부터 선택하는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the Z or triple agent is a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 77 and 96 do it with
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 파네소이드 X 수용체 작용제는 카페스톨(Cafestol), 케노데옥시콜린산(Chenodeoxycholic acid), 오베티콜린산(Obeticholic acid), 펙사라민(Fexaramine), GW 4064, PX104, 6E-CDCA(6-ethyl-chedeoxycholic acid), AKN-083, 트로피펙서(Tropifexor), 킬로펙서(Cilofexor), EDP-305, AGN-242266, AGN-242256, EP-024297, RDX-023, BWL-200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, PX20606, EYP-001, TERN-101, TC-100 및 INT-2228으로 이루어진 군에서 선택하는 하나 이상인 것을 특징으로 한다.As a pharmaceutical composition according to any one of the preceding embodiments, the farnesoid X receptor agonist is cafestol (Cafestol), chenodeoxycholic acid, obeticholic acid, fexaramine ( Fexaramine), GW 4064, PX104, 6E-CDCA (6-ethyl-chedeoxycholic acid), AKN-083, Tropifexor, Cilofexor, EDP-305, AGN-242266, AGN-242256, EP- 024297, RDX-023, BWL-200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, PX20606, It is characterized in that at least one selected from the group consisting of EYP-001, TERN-101, TC-100 and INT-2228.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 약학적 조성물은 아세틸조효소 A 카르복실라제 억제제(acetyl-Coenzyme A carboxylase inhibitor)와도 병용하는 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the pharmaceutical composition is also used in combination with an acetyl-Coenzyme A carboxylase inhibitor.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 아세틸조효소 A 카르복실라제 억제제는 CP-640186,(4-피페리디닐)-피페라진 유도체((4-piperidinyl)-piperazine derivatives), 1,4-치환 사이클로헥산 유도체(1,4-disubstituted cyclohexane derivatives), 스피로크로마논 유도체(Spirochromanone derivatives), 스피로락탐 유도체(Spirolactam derivatives), 스피로디아민 유도체(Spirodiamine derivatives), 스피로펜타실아미드 유도체(Spiropentacylamide derivatives), 슈도펩타이드 피롤리딘다이온 유도체(Pseudopeptide pyrrolidinedione derivatives), 매크로사이클릭 폴리케톤 유도체(Macrocyclic polyketone derivatives), 피르소코스타트 (firsocostat)를 포함하는 티오펜 피리미돈 유도체(Thiophene pyrimidone derivatives), 아미노-옥사졸 유도체(Amino-oxazole derivatives), 아조벤지미다졸 유도체(Azobenzimidazole derivatives) 및 PF-05221304로 이루어진 군에서 선택하는 하나 이상인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the acetyl coenzyme A carboxylase inhibitor is CP-640186, (4-piperidinyl)-piperazine derivatives ((4-piperidinyl)-piperazine derivatives), 1, 4-substituted cyclohexane derivatives, spirochromanone derivatives, spirolactam derivatives, spirodiamine derivatives, spiropentacylamide derivatives , Pseudopeptide pyrrolidinedione derivatives, Macrocyclic polyketone derivatives, Thiophene pyrimidone derivatives including firsocostat, amino-oxazole It is characterized in that at least one selected from the group consisting of amino-oxazole derivatives, azobenzimidazole derivatives and PF-05221304.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 간 질환은 비알코올성 지방간 질환(non-alcoholic fatty liver disease) 또는 담즙정체성 간질환인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the liver disease is non-alcoholic fatty liver disease or cholestatic liver disease.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 비알코올성 지방간 질환은 단순 지방증, 간 염증, 비알코올성 지방간, 비알코올성 지방간염, 간경변, 간 섬유증, 간부전, 및 간암으로 이루어진 군에서 선택하는 적어도 하나의 질환인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the nonalcoholic fatty liver disease is at least selected from the group consisting of steatosis simplex, liver inflammation, nonalcoholic fatty liver, nonalcoholic steatohepatitis, cirrhosis, liver fibrosis, liver failure, and liver cancer. It is characterized as a single disease.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 담즙 정체성 간질환은 원발성 담즙성 경변증 (Primary biliary cirrhosis), 원발성 경화성 담관염(primary sclerosing cholangitis) 및 이들의 조합으로 이루어진 군에서 선택하는 어느 하나인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the cholestatic liver disease is any one selected from the group consisting of primary biliary cirrhosis, primary sclerosing cholangitis, and combinations thereof. characterized in that
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 비알코올성 지방간 질환은 간 염증, 비알코올성 지방간염과 간 섬유증으로 이루어진 군에서 선택하는 적어도 하나의 질환인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the nonalcoholic fatty liver disease is at least one disease selected from the group consisting of liver inflammation, nonalcoholic steatohepatitis and liver fibrosis.
본 발명을 구현하는 또 하나의 양태는 (i) 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 갖는 물질 또는 이의 지속형 결합체, 및 (ii) FXR 작용제 (farnesoid X receptor agonist)를 포함하는, 조합물이다.Another aspect embodying the present invention is (i) a substance having activity against glucagon receptors, GLP-1 (Glucagon-like peptide-1) receptors, and GIP (Glucose-dependent insuliontropic polypeptide) receptors or long-acting conjugates thereof , and (ii) a farnesoid X receptor agonist.
하나의 구체예로서, 상기 조합물은 (iii) ACC 억제제 (acetyla-CoA carboxylase inhibitor)를 추가로 포함하는 것을 특징으로 한다.In one embodiment, the combination further comprises (iii) an acetyla-CoA carboxylase inhibitor.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질은 상기에서 설명한 일반식 1로 표시되는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor is a peptide comprising the amino acid sequence represented by the general formula 1 described above. do.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 일반식 1에서,A combination according to any one of the preceding embodiments, wherein in the general formula 1,
Xaa14는 류신 또는 메티오닌이며, Xaa14 is leucine or methionine,
Xaa15는 시스테인, 아스파르트산, 또는 류신인 것을 특징으로 한다.Xaa15 is characterized in that it is cysteine, aspartic acid, or leucine.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질은 상기에서 설명한 일반식 2로 표시되는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다The combination according to any one of the preceding embodiments, wherein the substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor is a peptide comprising the amino acid sequence represented by the general formula 2 described above. do
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 일반식 1에서,A combination according to any one of the preceding embodiments, wherein in the general formula 1,
Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이고,Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
Xaa7은 트레오닌이며,Xaa7 is threonine,
Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa10 is tyrosine, cysteine, or valine,
Xaa12는 리신 또는 이소류신이며, Xaa12 is lysine or isoleucine,
Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa13 is tyrosine, alanine, or cysteine,
Xaa14는 류신 또는 메티오닌이며,Xaa14 is leucine or methionine,
Xaa15는 시스테인 또는 아스파르트산이고,Xaa15 is cysteine or aspartic acid,
Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 또는 리신이며,Xaa17 is glutamine, arginine, isoleucine, cysteine, or lysine;
Xaa18은 알라닌, 아르기닌, 또는 히스티딘이고,Xaa18 is alanine, arginine, or histidine;
Xaa19는 알라닌, 글루타민, 또는 시스테인이며,Xaa19 is alanine, glutamine, or cysteine,
Xaa20은 리신 또는 글루타민이고,Xaa20 is lysine or glutamine,
Xaa21은 글루탐산, 시스테인, 또는 아스파르트산이며,Xaa21 is glutamic acid, cysteine, or aspartic acid;
Xaa23은 발린이고,Xaa23 is valine,
Xaa24는 알라닌, 글루타민, 시스테인, 아스파라긴, 또는 아스파르트산이며,Xaa24 is alanine, glutamine, cysteine, asparagine, or aspartic acid;
Xaa27은 류신 또는 리신인 것을 특징으로 한다.Xaa27 is characterized in that it is leucine or lysine.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 일반식 2에서,A combination according to any one of the preceding embodiments, wherein in the general formula 2,
Xaa13은 알라닌, 티로신, 또는 시스테인이고,Xaa13 is alanine, tyrosine, or cysteine;
Xaa15는 아스파르트산 또는 글루탐산이며,Xaa15 is aspartic acid or glutamic acid,
Xaa17은 글루타민, 아르기닌, 시스테인, 또는 리신이고,Xaa17 is glutamine, arginine, cysteine, or lysine;
Xaa18은 알라닌, 아르기닌, 또는 히스티딘이며,Xaa18 is alanine, arginine, or histidine;
Xaa21은 시스테인, 글루탐산, 글루타민, 또는 아스파르트산이고,Xaa21 is cysteine, glutamic acid, glutamine, or aspartic acid;
Xaa23은 이소류신 또는 발린이며,Xaa23 is isoleucine or valine,
Xaa24는 시스테인, 글루타민, 또는 아스파라긴이고, Xaa24 is cysteine, glutamine, or asparagine;
Xaa28은 시스테인, 아스파라긴, 또는 아스파르트산이며,Xaa28 is cysteine, asparagine, or aspartic acid;
Xaa29는 글루타민, 시스테인, 또는 히스티딘이고,Xaa29 is glutamine, cysteine, or histidine;
Xaa30은 시스테인, 리신, 또는 히스티딘인 것을 특징으로 한다.Xaa30 is characterized in that it is cysteine, lysine, or histidine.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 일반식 1에서,A combination according to any one of the preceding embodiments, wherein in the general formula 1,
Xaa2는 알파-메틸-글루탐산 또는 Aib이고,Xaa2 is alpha-methyl-glutamic acid or Aib;
Xaa7은 트레오닌이며,Xaa7 is threonine,
Xaa10은 티로신 또는 시스테인이고,Xaa10 is tyrosine or cysteine,
Xaa12는 리신 또는 이소류신이며, Xaa12 is lysine or isoleucine,
Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa13 is tyrosine, alanine, or cysteine,
Xaa14는 류신 또는 메티오닌이며,Xaa14 is leucine or methionine,
Xaa15는 시스테인 또는 아스파르트산이고,Xaa15 is cysteine or aspartic acid,
Xaa16은 글루탐산이며,Xaa16 is glutamic acid,
Xaa17은 아르기닌, 이소류신, 시스테인, 또는 리신이고,Xaa17 is arginine, isoleucine, cysteine, or lysine;
Xaa18은 알라닌, 아르기닌, 또는 히스티딘이며,Xaa18 is alanine, arginine, or histidine;
Xaa19는 알라닌, 글루타민, 또는 시스테인이고,Xaa19 is alanine, glutamine, or cysteine;
Xaa20은 리신 또는 글루타민이며,Xaa20 is lysine or glutamine,
Xaa21은 글루탐산 또는 아스파르트산이고,Xaa21 is glutamic acid or aspartic acid,
Xaa23은 발린이며,Xaa23 is valine,
Xaa24는 글루타민, 아스파라긴, 또는 아스파르트산이고,Xaa24 is glutamine, asparagine, or aspartic acid;
Xaa27은 류신이며,Xaa27 is leucine,
Xaa28은 시스테인, 알라닌, 아스파라긴, 또는 아스파르트산인 것을 특징으로 한다.Xaa28 is characterized in that it is cysteine, alanine, asparagine, or aspartic acid.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 일반식 1에서,A combination according to any one of the preceding embodiments, wherein in the general formula 1,
Xaa1은 히스티딘 또는 4-이미다조아세틸이고,Xaa1 is histidine or 4-imidazoacetyl;
Xaa2는 알파-메틸-글루탐산 또는 Aib이며,Xaa2 is alpha-methyl-glutamic acid or Aib;
Xaa3은 글루타민이고, Xaa3 is glutamine,
Xaa7은 트레오닌이며,Xaa7 is threonine,
Xaa10은 티로신이고, Xaa10 is tyrosine,
Xaa12는 이소류신이며, Xaa12 is isoleucine,
Xaa13은 알라닌 또는 시스테인이고, Xaa13 is alanine or cysteine,
Xaa14는 메티오닌이며,Xaa14 is methionine,
Xaa15는 아스파르트산이고,Xaa15 is aspartic acid,
Xaa16은 글루탐산이며,Xaa16 is glutamic acid,
Xaa17은 이소류신 또는 리신이고,Xaa17 is isoleucine or lysine,
Xaa18은 알라닌 또는 히스티딘이며,Xaa18 is alanine or histidine,
Xaa19는 글루타민 또는 시스테인이고,Xaa19 is glutamine or cysteine,
Xaa20은 리신이며,Xaa20 is lysine,
Xaa21은 아스파르트산이고,Xaa21 is aspartic acid,
Xaa23은 발린이며,Xaa23 is valine,
Xaa24는 아스파라긴이고,Xaa24 is asparagine,
Xaa27은 류신이며,Xaa27 is leucine,
Xaa28은 알라닌 또는 아스파라긴이고,Xaa28 is alanine or asparagine,
Xaa29는 글루타민 또는 트레오닌이며,Xaa29 is glutamine or threonine,
Xaa30은 시스테인 또는 리신이거나, 부존재하는 것을 특징으로 한다.Xaa30 is characterized in that it is cysteine or lysine, or is absent.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질은 상기에서 설명한 일반식 3의 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor is a peptide comprising the amino acid sequence of Formula 3 as described above.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 일반식 1에서,A combination according to any one of the preceding embodiments, wherein in the general formula 1,
상기 R1은 시스테인, GKKNDWKHNIT (서열번호 106), CSSGQPPPS (서열번호 109), GPSSGAPPPS (서열번호 110), GPSSGAPPPSC (서열번호 111), PSSGAPPPS (서열번호 112), PSSGAPPPSG (서열번호 113), PSSGAPPPSHG (서열번호 114), PSSGAPPPSS (서열번호 115), PSSGQPPPS (서열번호 116), 또는 PSSGQPPPSC (서열번호 117)이거나, 부존재하는 것을 특징으로 한다.R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), CSSGQPPPS (SEQ ID NO: 109), GPSSGAPPPS (SEQ ID NO: 110), GPSSGAPPPSC (SEQ ID NO: 111), PSSGAPPPS (SEQ ID NO: 112), PSSGAPPPSG (SEQ ID NO: 113), PSSGAPPPSHG (SEQ ID NO: 113) 114), PSSGAPPPSS (SEQ ID NO: 115), PSSGQPPPS (SEQ ID NO: 116), or PSSGQPPPSC (SEQ ID NO: 117), or is characterized in that it is absent.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 펩타이드는 상기 일반식에서 N-말단으로부터 16번 아미노산과 20번 아미노산은 서로 고리를 형성하는 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the peptide is characterized in that in the general formula, amino acids 16 and 20 from the N-terminus form a ring with each other.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 펩타이드는 이의 C-말단이 아미드화되거나 또는 자유 카르복실기 (-COOH)를 갖는 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the peptide is characterized in that its C-terminus is amidated or has a free carboxyl group (-COOH).
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질은 서열번호: 1 내지 102로 이루어진 군으로부터 선택된 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor is a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102 characterized.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 FXR 작용제는 카페스톨 (Cafestol), 케노데옥시콜산 (Chenodeoxycholic acid), 오베티콜산 (Obeticholic acid), 펙사라민 (Fexaramine), GW 4064, PX104, 6E-CDCA (6-ethyl-chedeoxycholic acid), AKN-083 및 트로피펙서 (Tropifexor), 킬로펙서 (Cilofexor), EDP-305 AGN-242266, AGN-242256, EP-024297, RDX-023, BWL-200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, PX20606, EYP-001, TERN-101, TC-100 및 INT-2228으로 이루어진 군에서 선택된 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the FXR agonist is Cafestol, Chenodeoxycholic acid, Obeticholic acid, Fexaramine, GW 4064, PX104 , 6E-CDCA (6-ethyl-chedeoxycholic acid), AKN-083 and Tropifexor, Cilofexor, EDP-305 AGN-242266, AGN-242256, EP-024297, RDX-023, BWL- 200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, PX20606, EYP-001, TERN-101, It is characterized in that it is selected from the group consisting of TC-100 and INT-2228.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 ACC 억제제는 CP-640186, (4-피페리디닐)-피페라진 유도체 ((4-piperidinyl)-piperazine derivatives), 1,4-치환 사이클로헥산 유도체 (1,4-disubstituted cyclohexane derivatives), 스피로크로마논 유도체 (Spirochromanone derivatives), 스피로락탐 유도체 (Spirolactam derivatives), 스피로디아민 유도체 (Spirodiamine derivatives), 스피로펜타실아미드 유도체 (Spiropentacylamide derivatives), 슈도펩타이드 피롤리딘다이온 유도체 (Pseudopeptide pyrrolidinedione derivatives), 매크로사이클릭 폴리케톤 유도체 (Macrocyclic polyketone derivatives), 피르소코스타트 (firsocostat)를 포함하는 티오펜 피리미돈 유도체 (Thiophene pyrimidone derivatives), 아미노-옥사졸 유도체 (Amino-oxazole derivatives), 아조벤지미다졸 유도체 (Azobenzimidazole derivatives) 및 PF-05221304로 이루어진 군에서 선택된 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the ACC inhibitor is CP-640186, (4-piperidinyl)-piperazine derivatives, 1,4-substituted cyclohexane derivatives (1,4-disubstituted cyclohexane derivatives), Spirochromanone derivatives, Spirolactam derivatives, Spirodiamine derivatives, Spiropentacylamide derivatives, Pseudopeptide pyrrolidine Ionic derivatives (Pseudopeptide pyrrolidinedione derivatives), Macrocyclic polyketone derivatives (Macrocyclic polyketone derivatives), Thiophene pyrimidone derivatives including firsocostat, Amino-oxazole derivatives ), azobenzimidazole derivatives, and PF-05221304.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 결합체는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질의 생체 내 반감기를 증가시킬 수 있는 생체적합성 물질이 결합된, 지속형 결합체의 형태인 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the conjugate is a glucagon receptor, a GLP-1 receptor, and a biocompatible substance capable of increasing the in vivo half-life of a substance having activity on the GIP receptor bound, long-acting It is characterized in that it is in the form of a conjugate.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 결합체는 상기에서 설명한 화학식 1로 표시되는 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the conjugate is represented by Formula 1 described above.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 화학식 1의 Fc는 면역글로불린 Fc 영역인 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the Fc of Formula 1 is an immunoglobulin Fc region.
앞선 구체예 중 어느 하나에 따른 약학적 조성물 또는 조합물로서, 상기 Fc 영역은 IgG Fc 영역인 것을 특징으로 한다.The pharmaceutical composition or combination according to any one of the preceding embodiments, wherein the Fc region is an IgG Fc region.
앞선 구체예 중 어느 하나에 따른 약학적 조성물 또는 조합물로서, 상기 Lx는 폴리에틸렌 글리콜인 것을 특징으로 한다.The pharmaceutical composition or combination according to any one of the preceding embodiments, wherein Lx is polyethylene glycol.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 조합물은 간 질환의 예방 또는 치료에 사용하기 위한 것을 특징으로 한다.A combination according to any one of the preceding embodiments, characterized in that the combination is for use in the prophylaxis or treatment of liver disease.
앞선 구체예 중 어느 하나에 따른 조합물로서, 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질은 서열번호: 1 내지 102로 이루어진 군으로부터 선택된 아미노산 서열을 포함하는 펩타이드이고, 이의 결합체는 상기 펩타이드와 면역글로불린 Fc 영역이 비펩타이드성 중합체인 링커를 통해 연결된 것을 특징으로 한다.The combination according to any one of the preceding embodiments, wherein the substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor is a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102, The conjugate thereof is characterized in that the peptide and the immunoglobulin Fc region are linked through a linker, which is a non-peptide polymer.
본 발명을 구현하는 다른 하나의 양태는 상기 조합물을 포함하는, 간 질환의 예방 또는 치료용 약학적 조성물이다.Another embodiment embodying the present invention is a pharmaceutical composition for preventing or treating liver disease, comprising the combination.
하나의 구체예로서, 상기 간 질환은 비알코올성 지방간 질환(non-alcoholic fatty liver disease) 또는 담즙정체성 간질환인 것을 특징으로 한다.In one embodiment, the liver disease is characterized in that non-alcoholic fatty liver disease (non-alcoholic fatty liver disease) or cholestatic liver disease.
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 비알코올성 지방간 질환은 단순 지방증, 간 염증, 비알코올성 지방간, 비알코올성 지방간염, 간경변, 간 섬유증, 간 부전 및 간암으로 이루어진 군에서 선택된 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the nonalcoholic fatty liver disease is selected from the group consisting of simple steatosis, liver inflammation, nonalcoholic fatty liver, nonalcoholic steatohepatitis, cirrhosis, liver fibrosis, liver failure and liver cancer. do it with
앞선 구체예 중 어느 하나에 따른 약학적 조성물로서, 상기 담즙 정체성 간질환은 원발성 담즙성 경변증, 원발성 경화성 담관염 및 이들의 조합으로 이루어진 군에서 선택하는 어느 하나인 것을 특징으로 한다.The pharmaceutical composition according to any one of the preceding embodiments, wherein the cholestatic liver disease is characterized in that it is any one selected from the group consisting of primary biliary cirrhosis, primary sclerosing cholangitis, and combinations thereof.
본 발명을 구현하는 다른 하나의 양태는 FXR 작용제 (farnesoid X receptor agonist)와 병용하는 것을 특징으로 하는 상기의 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 갖는 물질 또는 이의 결합체를 포함하는, 간 질환의 예방 또는 치료용 약학적 조성물이다.Another aspect of implementing the present invention is the above glucagon receptor, GLP-1 (Glucagon-like peptide-1) receptor, and GIP (glucose-dependent insuliontropic), characterized in that it is used in combination with an FXR agonist (farnesoid X receptor agonist). Polypeptide) is a pharmaceutical composition for preventing or treating liver disease, including a substance having activity on the receptor or a conjugate thereof.
하나의 구체예로서, 상기 조성물은 추가로 ACC 억제제 (acetyla-CoA carboxylase inhibitor)와 병용하는 것을 특징으로 한다.In one embodiment, the composition is characterized in that it is further used in combination with an ACC inhibitor (acetyla-CoA carboxylase inhibitor).
본 발명을 구현하는 다른 하나의 양태는 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 또는 이의 지속형 결합체, 및 (ii) FXR 작용제를 포함하는, 간 질환의 예방 또는 치료를 위한 약학적 키트이다.Another aspect embodying the present invention is (i) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor, or a long-acting conjugate thereof, and (ii) an FXR agonist, the prevention of liver disease or a pharmaceutical kit for treatment.
하나의 구체예로서, 상기 키트는 (iii) ACC 억제제를 추가로 포함하는 것을특징으로 한다.In one embodiment, the kit is characterized in that it further comprises (iii) an ACC inhibitor.
본 발명을 구현하는 다른 하나의 양태는 상기 조합물, 상기 간 질환의 치료 또는 예방을 위한 약학적 조성물, 또는 상기 약학적 키트를 이를 필요로 하는 개체에 투여 및/또는 사용하는 단계를 포함하는 간 질환의 예방 또는 치료 방법이다.Another embodiment embodying the present invention is a liver comprising the step of administering and/or using the combination, a pharmaceutical composition for the treatment or prevention of liver disease, or the pharmaceutical kit to an individual in need thereof A method for preventing or treating a disease.
본 발명을 구현하는 다른 하나의 양태는 상기 조합물, 약학적 조성물, 또는 약학적 키트의 간 질환의 예방 또는 치료 용도 및/또는 간 질환의 예방 또는 치료를 위한 약제의 제조를 위한 용도이다.Another aspect embodying the present invention is the use of the combination, pharmaceutical composition, or pharmaceutical kit for the prophylaxis or treatment of liver disease and/or use for the manufacture of a medicament for the prophylaxis or treatment of liver disease.
이하에서는, 본 발명을 더욱 상세히 설명한다.Hereinafter, the present invention will be described in more detail.
한편, 본원에서 개시되는 각각의 설명 및 실시형태는 각각의 다른 설명 및 실시 형태에도 적용될 수 있다. 즉, 본원에서 개시된 다양한 요소들의 모든 조합이 본 발명의 범주에 속한다. 또한, 하기 기술되는 구체적인 서술에 의하여 본 발명의 범주가 제한된다고 할 수 없다.On the other hand, each description and embodiment disclosed herein may be applied to each other description and embodiment. That is, all combinations of the various elements disclosed herein are within the scope of the present invention. In addition, it cannot be said that the scope of the present invention is limited by the specific descriptions described below.
또한, 당해 기술분야의 통상의 지식을 가진 자는 통상의 실험만을 사용하여 본 출원에 기재된 본 발명의 특정 양태에 대한 다수의 등가물을 인지하거나 확인할 수 있다. 또한, 이러한 등가물은 본 발명에 포함되는 것으로 의도된다. In addition, those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. Also, such equivalents are intended to be encompassed by the present invention.
본 명세서 전반을 통하여, 천연적으로 존재하는 아미노산에 대한 통상의 1문자 및 3문자 코드가 사용될 뿐만 아니라 Aib(2-아미노이소부티르산, 2-aminoisobutyric acid), Sar(N-methylglycine) 등과 같은 다른 아미노산에 대해 일반적으로 허용되는 3문자 코드가 사용된다. 또한 본 명세서에서 약어로 언급된 아미노산은 IUPAC-IUB 명명법에 따라 기재되었다. Throughout this specification, common one-letter and three-letter codes for naturally occurring amino acids are used as well as other amino acids such as Aib (2-aminoisobutyric acid, 2-aminoisobutyric acid), Sar (N-methylglycine), etc. The generally accepted three-letter code for Amino acids referred to by abbreviation herein are also described according to the IUPAC-IUB nomenclature.
알라닌 Ala, A 아르기닌 Arg, RAlanine Ala, A Arginine Arg, R
아스파라긴 Asn, N 아스파르트산 Asp, DAsparagine Asn, N Aspartic Acid Asp, D
시스테인 Cys, C 글루탐산 Glu, ECysteine Cys, C Glutamate Glu, E
글루타민 Gln, Q 글리신 Gly, GGlutamine Gln, Q Glycine Gly, G
히스티딘 His, H 이소류신 Ile, IHistidine His, H Isoleucine Ile, I
류신 Leu, L 리신 Lys, KLeucine Leu, L Lysine Lys, K
메티오닌 Met, M 페닐알라닌 Phe, FMethionine Met, M Phenylalanine Phe, F
프롤린 Pro, P 세린 Ser, SProline Pro, P Serine Ser, S
트레오닌 Thr, T 트립토판 Trp, WThreonine Thr, T Tryptophan Trp, W
티로신 Tyr, Y 발린 Val, VTyrosine Tyr, Y Valine Val, V
본 발명을 구현하는 하나의 양태는 3중 작용성 지속형 결합체(triple agonistic long-acting conjugate) 또는 3중 작용제를 포함하는 간 질환의 치료 또는 예방을 위한 약학적 조성물로서, 상기 약학적 조성물은 파네소이드 X 수용체 작용제(farnesoid X receptor agonist, FXR 작용제)와 병용되는 것을 특징하는 약학적 조성물을 제공한다.One aspect embodying the present invention is a pharmaceutical composition for the treatment or prevention of liver disease comprising a triple agonistic long-acting conjugate or a triple agonist, wherein the pharmaceutical composition is Provided is a pharmaceutical composition characterized in that it is used in combination with a farnesoid X receptor agonist (FXR agonist).
상기 "3중 작용성 지속형 결합체"는 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 갖는, 펩타이드에 이의 생체 내 반감기를 증가시키는 생체 적합성 물질부가 결합된 지속형 결합체를 의미한다. 본 명세서에서 상기 생체적합성 물질은 캐리어와 혼용될 수 있다. 상기 "글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (또는 펩타이드)"는 "3중 작용제 (triple agonis, trigonal agonist)" 또는 “삼중 활성체”로 지칭될 수 있다.The "triple-acting long-acting conjugate" is a glucagon receptor, GLP-1 (Glucagon-like peptide-1) receptor, and GIP (Glucose-dependent insuliontropic polypeptide) having activity on the receptor, the peptide having its in vivo half-life It refers to a long-acting conjugate to which the increasing biocompatible material part is bound. In the present specification, the biocompatible material may be mixed with a carrier. The "substance (or peptide) having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor" may be referred to as a "triple agonis, trigonal agonist" or "triple activator".
본 발명에서 상기 펩타이드의 결합체인 "3중 작용성 지속형 결합체"는 캐리어가 결합되지 않은 상기 펩타이드에 비하여 증가된 효력의 지속성을 나타낼 수 있으며, 본 발명에서는 이러한 결합체를 "지속형 결합체"로 지칭하고 "결합체", "삼중 활성체의 지속형 결합체", 또는 "글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질의 결합체"로 혼용되어 사용될 수 있다.In the present invention, the "triple-acting long-acting conjugate", which is a conjugate of the peptide, may exhibit increased durability of potency compared to the peptide to which a carrier is not bound, and in the present invention, such a conjugate is referred to as a "long-acting conjugate" and "conjugate", "long-acting conjugate of triple activator", or "conjugate of substances having activity against glucagon receptor, GLP-1 receptor, and GIP receptor" may be used interchangeably.
본 발명의 하나의 구체예에서, 상기 약학적 조성물은 약학적 유효량의 3중 작용성 지속형 결합체 또는 3중 작용제와 약학적으로 허용되는 부형제를 함유하는 간 질환의 치료 또는 예방을 위한 약학적 조성물로서,In one embodiment of the present invention, the pharmaceutical composition is a pharmaceutical composition for the treatment or prevention of liver disease containing a pharmaceutically effective amount of a triple-acting long-acting conjugate or a triple agent and a pharmaceutically acceptable excipient. as,
상기 약학적 조성물은 파네소이드 X 수용체 작용제와 병용되는 것을 특징으로 한다.The pharmaceutical composition is characterized in that it is used in combination with a farnesoid X receptor agonist.
상기 "3중 작용성 지속형 결합체"는 하기 화학식 1로 표시되는 물질일 수 있으나, 이에 제한되지 않는다.The "tri-functional long-acting conjugate" may be a material represented by the following formula (1), but is not limited thereto.
[화학식 1][Formula 1]
Z - Lx - FcZ - Lx - Fc
이 때 Lx는 에틸렌글리콜 반복 단위를 함유하는 링커이며, x는 0 또는 자연수이고,In this case, Lx is a linker containing an ethylene glycol repeating unit, and x is 0 or a natural number,
Fc는 면역글로불린 Fc 영역이고, Fc is an immunoglobulin Fc region,
-는 Lx와 Z 사이, Fc와 Lx 사이의 각각에 공유결합 연결을 나타내며,- represents a covalent linkage between Lx and Z, between Fc and Lx, respectively,
Z 또는 3중 작용제는 하기 일반식 1로 표시되는 아미노산 서열을 포함하는 펩타이드이고, Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 1,
Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1(일반식 1, 서열번호 103)Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp23 Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (Formula 1, SEQ ID NO: 103)
상기 일반식 1에서,In the above general formula 1,
Xaa1은 히스티딘(His, H), 4-이미다조아세틸(CA) 또는 티로신(Tyr, Y)이고, Xaa1 is histidine (His, H), 4-imidazoacetyl (CA) or tyrosine (Tyr, Y),
Xaa2는 글리신(Gly, G), 알파-메틸-글루탐산 또는 2-아미노이소부티르산(2-aminoisobutyric acid, Aib)이며,Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid or 2-aminoisobutyric acid (Aib),
Xaa3은 글루탐산(Glu, E) 또는 글루타민(Gln, Q)이고, Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q),
Xaa7은 트레오닌(Thr, T) 또는 이소류신(Ile, I)이며,Xaa7 is threonine (Thr, T) or isoleucine (Ile, I),
Xaa10은 류신(Leu, L), 티로신(Tyr, Y), 리신(Lys, K), 시스테인(Cys, C) 또는 발린(Val, V)이고, Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C) or valine (Val, V);
Xaa12는 리신(Lys, K), 세린(Ser, S) 또는 이소류신(Ile, I)이며, Xaa12 is lysine (Lys, K), serine (Ser, S) or isoleucine (Ile, I);
Xaa13은 글루타민(Gln, Q), 티로신(Tyr, Y), 알라닌(Ala, A) 또는 시스테인(Cys, C)이고, Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A) or cysteine (Cys, C);
Xaa14는 류신(Leu, L), 메티오닌(Met, M) 또는 티로신(Tyr, Y)이며,Xaa14 is leucine (Leu, L), methionine (Met, M) or tyrosine (Tyr, Y);
Xaa15는 시스테인(Cys, C), 아스파르트산(Asp, D), 글루탐산(Glu, E) 또는 류신(Leu, L)이며,Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E) or leucine (Leu, L),
Xaa16은 글리신(Gly, G), 글루탐산(Glu, E) 또는 세린(Ser, S)이고, Xaa16 is glycine (Gly, G), glutamic acid (Glu, E) or serine (Ser, S);
Xaa17은 글루타민(Gln, Q), 아르기닌(Arg, R), 이소류신(Ile, I), 글루탐산(Glu, E), 시스테인(Cys, C) 또는 리신(Lys, K)이며,Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C) or lysine (Lys, K);
Xaa18은 알라닌(Ala, A), 글루타민(Gln, Q), 아르기닌(Arg, R) 또는 히스티딘(His, H)이고,Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R) or histidine (His, H);
Xaa19는 알라닌(Ala, A), 글루타민(Gln, Q), 시스테인(Cys, C) 또는 발린(Val, V)이며,Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C) or valine (Val, V);
Xaa20은 리신(Lys, K), 글루타민(Gln, Q) 또는 아르기닌(Arg, R)이고,Xaa20 is lysine (Lys, K), glutamine (Gln, Q) or arginine (Arg, R);
Xaa21은 글루탐산(Glu, E), 글루타민(Gln, Q), 류신(Leu, L), 시스테인(Cys, C) 또는 아스파르트산(Asp, D)이며,Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C) or aspartic acid (Asp, D);
Xaa23은 이소류신(Ile, I) 또는 발린(Val, V)이고,Xaa23 is isoleucine (Ile, I) or valine (Val, V),
Xaa24는 알라닌(Ala, A), 글루타민(Gln, Q), 시스테인(Cys, C), 아스파라긴(Asn, N), 아스파르트산(Asp, D) 또는 글루탐산(Glu, E)이며,Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D) or glutamic acid (Glu, E),
Xaa27은 발린(Val, V), 류신(Leu, L) 또는 리신(Lys, K) 이고,Xaa27 is valine (Val, V), leucine (Leu, L) or lysine (Lys, K);
Xaa28은 시스테인(Cys, C), 리신(Lys, K), 알라닌(Ala, A), 아스파라긴(Asn, N) 또는 아스파르트산(Asp, D)이며,Xaa28 is cysteine (Cys, C), lysine (Lys, K), alanine (Ala, A), asparagine (Asn, N) or aspartic acid (Asp, D);
Xaa29는 시스테인(Cys, C), 글리신(Gly, G), 글루타민(Gln, Q), 트레오닌(Thr, T), 글루탐산(Glu, E) 또는 히스티딘(His, H)이고,Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q), threonine (Thr, T), glutamic acid (Glu, E) or histidine (His, H);
Xaa30은 시스테인(Cys, C), 글리신(Gly, G), 리신(Lys, K) 또는 히스티딘(His, H)이거나, 부존재하며,Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K) or histidine (His, H), or is absent;
R1은 시스테인(Cys, C), GKKNDWKHNIT(서열번호 106), m-SSGAPPPS-n(서열번호 107) 또는 m-SSGQPPPS-n(서열번호 108)이거나, 부존재하며, R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107) or m-SSGQPPPS-n (SEQ ID NO: 108), or is absent;
m은 -Cys-, -Pro- 또는 -Gly-Pro-이고, m is -Cys-, -Pro- or -Gly-Pro-;
n은 부존재하거나 -Cys-, -Gly-, -Ser- 또는 -His-Gly-이다.n is absent or is -Cys-, -Gly-, -Ser- or -His-Gly-.
상기 일반식 1은 서열번호 12일 경우 Xaa27은 예외적으로 메티오닌 (Met, M)일 수 있다.In the case of SEQ ID NO: 12 in Formula 1 above, Xaa27 may be exceptionally methionine (Met, M).
상기 약학적 조성물은 아세틸조효소 A 카르복실라제 억제제(acetyl-Coenzyme A carboxylase inhibitor, ACC 억제제)와도 병용하는 것일 수 있다.The pharmaceutical composition may be used in combination with an acetyl-Coenzyme A carboxylase inhibitor (ACC inhibitor).
본 발명의 또 하나의 양태는 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 또는 이의 지속형 결합체, 및 (ii) FXR 작용제 (farnesoid X receptor agonist)를 병용하는 치료학적 용도를 제공한다.Another aspect of the present invention is (i) a substance having activity on glucagon receptor, GLP-1 receptor, and GIP receptor or a long-acting conjugate thereof, and (ii) FXR agonist (farnesoid X receptor agonist) in combination treatment provide academic use.
또 하나의 구체예는 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 또는 이의 지속형 결합체, (ii) FXR 작용제, 및 (iii) ACC 억제제 (acetyla-CoA carboxylase inhibitor)를 병용하는 치료학적 용도를 제공한다.Another embodiment includes (i) a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors or long-acting conjugates thereof, (ii) FXR agonists, and (iii) ACC inhibitors (acetyla-CoA carboxylase inhibitors) ) provides a therapeutic use in combination.
또 하나의 구체예는 (i) 3중 작용성 지속형 결합체 또는 3중 작용제 및 (ii) FXR 작용제를 병용하는 간 질환의 예방 또는 치료용 약학적 조성물을 제공한다.Another embodiment provides a pharmaceutical composition for preventing or treating liver disease using (i) a triple-acting long-acting conjugate or a triple agonist and (ii) an FXR agonist.
또 하나의 구체예는 (i) 3중 작용성 지속형 결합체 또는 3중 작용제, (ii) FXR 작용제, 및 (iii) ACC 억제제를 병용하는 간 질환의 예방 또는 치료용 약학적 조성물을 제공한다.Another embodiment provides a pharmaceutical composition for the prophylaxis or treatment of liver disease using (i) a triple acting long-acting conjugate or triple agent, (ii) an FXR agonist, and (iii) an ACC inhibitor.
구체적으로, 본 발명의 일 양태는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체); 및 FXR 작용제를 포함하는 조합물, 약학적 조성물, 또는 키트를 제공한다. Specifically, one aspect of the present invention relates to a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and a combination, pharmaceutical composition, or kit comprising an FXR agonist.
일 구체예로, 상기 3중 작용제 또는 이의 지속형 결합체; 및 FXR 작용제를 포함하는 조합물, 약학적 조성물, 또는 키트는 간 질환의 예방 또는 치료용 조합물, 약학적 조성물, 또는 키트일 수 있다. In one embodiment, the triple agonist or a long-acting conjugate thereof; And the combination, pharmaceutical composition, or kit comprising a FXR agonist may be a combination, pharmaceutical composition, or kit for the prophylaxis or treatment of liver disease.
상기 조합물은 ACC 억제제를 추가로 포함할 수 있으며, 이 경우 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 결합체 (3중 작용성 지속형 결합체); FXR 작용제; 및 ACC 억제제를 포함하는 조합물, 약학적 조성물, 또는 키트를 제공한다. 일 구체예로, 상기 3중 작용제 또는 이의 지속형 결합체; FXR 작용제; 및 ACC 억제제를 포함하는 조합물, 약학적 조성물, 또는 키트는 간 질환의 예방 또는 치료용 조합물, 약학적 조성물, 또는 키트일 수 있다.The combination may further comprise an ACC inhibitor, in this case, a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate) ; FXR agonists; and an ACC inhibitor. In one embodiment, the triple agonist or a long-acting conjugate thereof; FXR agonists; And the combination, pharmaceutical composition, or kit comprising an ACC inhibitor may be a combination, pharmaceutical composition, or kit for the prophylaxis or treatment of liver disease.
본 발명에서 용어 "조합물(combination)"은 (a) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체); 및 (b) FXR 작용제, 또는 (b') FXR 작용제 및 ACC 억제제의 병용 투여 용도를 갖는 것으로, "병용 용도(combined used)"와 같은 의미로 이해될 수 있다. 이는 3중 작용성 지속형 결합체 또는 3중 작용제와 FXR 작용제가 병용되는 것을 특징으로 하는 약학적 조성물이나, 상기 약학적 조성물에 ACC 억제제가 추가로 병용되는 조성물 형태도 포함하나, 이에 제한되지 않는다. 본원에서 (a) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체); 및 (b) FXR 작용제의 병용 투여는 "복합 병용 투여" 또는 "복합 투여"로 혼용될 수 있으며, (a) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 결합체 (3중 작용성 지속형 결합체); 및 (b') FXR 작용제 및 ACC 억제제의 병용 투여는 "삼중 복합 병용 투여", "삼중 복합 투여"로 혼용될 수 있다.In the present invention, the term "combination" refers to (a) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) ); and (b) an FXR agonist, or (b') an FXR agonist and an ACC inhibitor. This includes, but is not limited to, a triple-acting long-acting conjugate or a pharmaceutical composition characterized in that a triple-agonist and an FXR agonist are used in combination, or a composition in which an ACC inhibitor is additionally used in combination with the pharmaceutical composition. As used herein, (a) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (b) concomitant administration of an FXR agonist may be used interchangeably as “combined administration” or “combined administration”, wherein (a) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) ) or a conjugate thereof (triple-acting long-acting conjugate); and (b') co-administration of an FXR agonist and an ACC inhibitor may be used interchangeably as "triple combined administration" and "triple combined administration".
상기 조합물 또는 약학적 조성물은The combination or pharmaceutical composition is
a) (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 (ii) FXR 작용제가 혼합된 하나의 혼합물(mixture)로 투여되는 것이거나, 또는 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), (ii) FXR 작용제, 및 (iii) ACC 억제제가 혼합된 하나의 혼합물(mixture)로 투여되는 것이거나; 또는a) (i) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) and (ii) an FXR agonist mixed It is administered as a mixture, or (i) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting type) conjugate), (ii) an FXR agonist, and (iii) an ACC inhibitor are administered in one mixture; or
b) (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 (ii) FXR 작용제가 분리된 형태로 투여되는 것이거나, 또는 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), (ii) FXR 작용제, 및 (iii) ACC 억제제가 분리된 형태로 투여되는 것일 수 있으나, 이에 제한되지 않는다. b) (i) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) and (ii) an FXR agonist isolated or (i) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), (ii) ) FXR agonist, and (iii) ACC inhibitor may be administered in separate form, but is not limited thereto.
글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제가 분리된 형태일 경우, 또는 GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제가 분리된 형태일 경우, 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제가 별개의 제제로 제제화되어 동시, 개별, 순차, 또는 역순으로 투여될 수 있는 것일 수 있다.A substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) and the FXR agonist in an isolated form, or GLP-1 When the receptor, and a substance having activity on the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist and an ACC inhibitor are in an isolated form, glucagon receptor, GLP-1 receptor , and substances having activity on the GIP receptor (triple agonist) or long-acting conjugates thereof (triple-acting long-acting conjugates) and FXR agonists; or a substance having activity on the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist and an ACC inhibitor are formulated in separate formulations, , sequentially, or in reverse order.
본 발명에서, "병용 투여", "병용되는" "병용하는"은 단지 동시의 투여를 의미하는 것뿐만이 아니라, a) (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 (ii) FXR 작용제; 또는 b) (i) GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), (ii) FXR 작용제 및 (iii) ACC 억제제가 개체에 함께 작용하여 각 물질 ((i), (ii) 및/또는 (iii))이 본연의 기능과 동등하거나 그 이상의 수준을 수행할 수 있는 투여 형태로 이해되어야 한다. 따라서, 본원에서, "병용"이란 용어가 사용될 경우 이는 동시, 개별, 순차, 또는 역순 투여를 나타내는 것으로 그 순서가 무제한 인 것으로 이해되어야 한다. 상기 투여가 순차, 역순 또는 개별적인 경우, 투여의 순서는 특별히 제한되지 않고 다만 2차 이상의 성분 투여의 간격은 상기 병용의 이로운 효과를 잃지 않도록 하는 것이어야 한다.In the present invention, "concomitant administration", "combination" and "in combination" do not only mean simultaneous administration, but also a) (i) having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor. a substance (a triple agonist) or a long-acting conjugate thereof (a triple-acting long-acting conjugate) and (ii) an FXR agonist; or b) (i) a GLP-1 receptor, and a substance having activity on the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), (ii) an FXR agonist and (iii) an ACC It is to be understood as a dosage form in which the inhibitors act together on the subject so that each of the substances ((i), (ii) and/or (iii)) performs at a level equal to or greater than its intended function. Accordingly, herein, when the term "combination" is used, it is to be understood that it refers to simultaneous, separate, sequential, or reverse administration, and the order is not limited. When the administration is sequential, reversed or individual, the order of administration is not particularly limited, provided that the interval between administration of the second or more components should be such that the beneficial effect of the combination is not lost.
본 발명에서 용어 "조합물을 포함하는 조성물(composition)"은 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체(3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제를 포함하는 조합물 그 자체이거나, 이를 포함하고, 치료학적 용도를 갖는 것일 수 있으나, 이에 제한되지 않는다. 그 예로, 간 질환의 예방 또는 치료 용도를 갖는 것일 수 있으나, 이에 제한되지 않는다. 본원에서 "조합물을 포함하는 조성물"은 "조성물"과 상호 교환적으로 사용될 수 있다.In the present invention, the term "composition comprising a combination" refers to a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple action lasting type conjugates) and FXR agonists; or a combination comprising a GLP-1 receptor, and a substance having activity on the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist and an ACC inhibitor, or and may have therapeutic uses, but is not limited thereto. For example, it may have a use for preventing or treating liver disease, but is not limited thereto. As used herein, “composition comprising a combination” may be used interchangeably with “composition”.
본 발명에 따른 조합물을 포함하는 조성물은 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제를 병용 투여하기 위한 것으로, 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제가 하나의 제제로 제제화된 것이거나, 또는 개별적으로 제제화된 것일 수 있다. 그 예로, 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체, FXR 작용제 및 ACC 억제제를 동시, 개별, 순차, 또는 역순으로 투여하는 것일 수 있으나, 이에 제한되지 않는다.The composition comprising the combination according to the present invention comprises a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) and an FXR agonist ; Or a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist and an ACC inhibitor for co-administration, substances having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonists) or long-acting conjugates thereof (triple-acting long-acting conjugates) and FXR agonists; or a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist and an ACC inhibitor are formulated in one formulation or may be individually formulated. For example, substances having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonists) or long-acting conjugates thereof (triple-acting long-acting conjugates) and FXR agonists; Or the GLP-1 receptor, and a substance having activity on the GIP receptor (triple agonist) or a long-acting conjugate thereof, an FXR agonist and an ACC inhibitor may be administered simultaneously, separately, sequentially, or in reverse order, but is not limited thereto does not
본 발명에서 용어 "키트"는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제를 병용 투여하기 위해, 본 발명에 따른 조합물 또는 조성물을 포함하는 것일 수 있다. 구체적으로, 본 발명에 따른 키트에는 하나의 제제로 제제화된 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제의 개별 제제를 포함하는 것일 수 있고, 두 물질, 또는 세 물질의 병용 투여에 필요한 물질을 추가적으로 포함하는 것일 수 있으나, 이에 제한되지 않는다. In the present invention, the term "kit" refers to a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) and an FXR agonist; or a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist and an ACC inhibitor, It may be one comprising the combination or composition according to the invention. Specifically, the kit according to the present invention includes a substance having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor formulated as one formulation (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) ) and FXR agonists; or substances having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonist) or long-acting conjugates thereof (triple-acting long-acting conjugates), FXR agonists and ACC inhibitors and may additionally include a material necessary for the combined administration of two substances or three substances, but is not limited thereto.
본 발명은 간 질환에 예방 또는 치료 효과를 갖는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체)에 FXR 작용제; 또는 FXR 작용제 및 ACC 억제제를 추가로 포함하여 병용하여 사용할 경우, 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 단독에 비해 간 질환의 예방 또는 치료 효과가 획기적으로 개선되는 것을 확인하여 상기 병용 사용 요법을 제공하게 되었다.The present invention relates to a substance having activity against glucagon receptors, GLP-1 receptors, and GIP receptors having a prophylactic or therapeutic effect on liver disease (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) FXR agent; Or when used in combination with an FXR agonist and an ACC inhibitor, a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting type) It was confirmed that the preventive or therapeutic effect of liver disease is dramatically improved compared to the conjugate) alone, and thus the combination therapy was provided.
상기 "글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는, 물질"은 "글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는, 펩타이드"일 수 있다. 본 발명에서 "3중 작용제 (triple agonist)" 또는 "삼중 활성체"로 혼용되어 사용될 수 있다. 상기 삼중 활성체는 상기 화학식 1의 일 구성요소인 Z일 수 있다.The “substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor” may be a “peptide having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor”. In the present invention, "triple agonist" or "triple active agent" may be used interchangeably. The triple activator may be Z, which is a component of Formula 1 above.
이러한 펩타이드는 글루카곤, GLP-1, 및 GIP 수용체에 대해 유의한 수준의 활성을 가지는 다양한 물질, 예컨대 다양한 펩타이드를 포함한다.Such peptides include various substances with significant levels of activity on glucagon, GLP-1, and GIP receptors, such as various peptides.
특별히 이에 제한되는 것은 아니나, 상기 글루카곤, GLP-1, 및 GIP 수용체에 대해 유의한 수준의 활성을 가지는 삼중 활성체는 글루카곤, GLP-1, 및 GIP 수용체 중 하나 또는 그 이상의 수용체, 구체적으로 둘 또는 그 이상의 수용체, 보다 구체적으로 세 개의 수용체 모두에 대해 in vitro 활성이 해당 수용체의 천연형 리간드 (천연형 글루카곤, 천연형 GLP-1, 및 천연형 GIP) 대비 약 0.001%이상, 약 0.01% 이상, 약 0.1% 이상, 약 1% 이상, 약 2% 이상, 약 3 % 이상, 약 4% 이상, 약 5% 이상, 약 6% 이상, 약 7% 이상, 약 8% 이상, 약 9% 이상, 약 10 % 이상, 약 20% 이상, 약 30% 이상, 약 40% 이상, 약 50 % 이상, 약 60% 이상, 약 70% 이상, 약 80% 이상, 약 90% 이상, 약 100%, 약 150% 이상, 약 200% 이상을 나타낼 수 있으나, 유의적으로 증가한 범위는 제한 없이 포함된다. Although not particularly limited thereto, the triple activator having a significant level of activity on the glucagon, GLP-1, and GIP receptors includes one or more receptors of glucagon, GLP-1, and GIP receptors, specifically two or more More specifically, the in vitro activity for all three receptors is about 0.001% or more, about 0.01% or more, compared to the native ligand (natural glucagon, native GLP-1, and native GIP) of the corresponding receptor; about 0.1% or more, about 1% or more, about 2% or more, about 3% or more, about 4% or more, about 5% or more, about 6% or more, about 7% or more, about 8% or more, about 9% or more, about 10% or more, about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60% or more, about 70% or more, about 80% or more, about 90% or more, about 100% or more, about It may represent 150% or more, about 200% or more, but a significantly increased range is included without limitation.
여기서, 수용체에 대한 활성은, 천연형 대비 수용체에 대한 in vitro 활성이 약 0.001%이상, 0.01% 이상, 0.1% 이상, 1% 이상, 2% 이상, 3 % 이상, 4% 이상, 5% 이상, 6% 이상, 7% 이상, 8% 이상, 9% 이상, 10 % 이상, 20% 이상, 30% 이상, 40% 이상, 50 % 이상, 60% 이상, 70% 이상, 80% 이상, 90 % 이상, 100% 이상, 약 200% 이상을 나타내는 경우를 예로 들 수 있다. 그러나, 이에 제한되는 것은 아니다. Here, the activity on the receptor is about 0.001% or more, 0.01% or more, 0.1% or more, 1% or more, 2% or more, 3% or more, 4% or more, 5% or more, the in vitro activity of the receptor compared to the native type. , 6% or more, 7% or more, 8% or more, 9% or more, 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, 90 % or more, 100% or more, or about 200% or more may be exemplified. However, the present invention is not limited thereto.
본 발명에서 용어, "약"은 ±0.5, ±0.4, ±0.3, ±0.2, ±0.1 등을 모두 포함하는 범위로, 약 이란 용어 뒤에 나오는 수치와 동등하거나 유사한 범위의 수치를 모두 포함하나, 이에 제한되지 않는다.In the present invention, the term “about” includes all values within a range including ±0.5, ±0.4, ±0.3, ±0.2, ±0.1, etc., and includes all values in a range equal to or similar to the value following the term about, but not limited
이러한 삼중 활성체의 in vitro 활성을 측정하는 방법은 본원 명세서의 실시예 3을 참조할 수 있으나, 특별히 이에 제한되는 것은 아니다. For a method of measuring the in vitro activity of such a triple activator, Example 3 of the present specification may be referred to, but is not particularly limited thereto.
한편, 상기 펩타이드는 하기 i) 내지 iii) 중 하나 이상, 둘 이상, 구체적으로 세 개의 활성을 보유하는 것, 구체적으로 유의한 활성을 보유하는 것을 특징으로 한다:On the other hand, the peptide is characterized in that it possesses one or more, two or more, specifically three activities of the following i) to iii), specifically, it possesses a significant activity:
i) GLP-1 수용체의 활성화; ii) 글루카곤 수용체의 활성화; 및 iii) GIP 수용체의 활성화.i) activation of the GLP-1 receptor; ii) activation of the glucagon receptor; and iii) activation of the GIP receptor.
여기서, 수용체를 활성화시킨다는 것은, 천연형 대비 수용체에 대한 in vitro 활성이 약 0.001%이상, 약 0.1% 이상, 약 1% 이상, 약 2% 이상, 약 3 % 이상, 약 4% 이상, 약 5% 이상, 약 6% 이상, 약 7% 이상, 약 8% 이상, 약 9% 이상, 약 10 % 이상, 약 20% 이상, 약 30% 이상, 약 40% 이상, 약 50 % 이상, 약 60% 이상, 약 70% 이상, 약 80% 이상, 약 90 % 이상, 약 100% 이상, 약 150% 이상, 약 200% 이상을 나타내는 경우를 예로 들 수 있다. 그러나, 이에 제한되는 것은 아니다.Here, activating the receptor means that the in vitro activity of the receptor compared to the native type is about 0.001% or more, about 0.1% or more, about 1% or more, about 2% or more, about 3% or more, about 4% or more, about 5 % or more, about 6% or more, about 7% or more, about 8% or more, about 9% or more, about 10% or more, about 20% or more, about 30% or more, about 40% or more, about 50% or more, about 60 % or more, about 70% or more, about 80% or more, about 90% or more, about 100% or more, about 150% or more, about 200% or more may be exemplified. However, the present invention is not limited thereto.
또한, 상기 펩타이드는 천연형 GLP-1, 천연형 글루카곤 및 천연형 GIP 중 어느 하나 대비 체내 반감기가 증가된 것일 수 있으나, 특별히 이에 제한되는 것은 아니다. In addition, the peptide may have an increased half-life in the body compared to any one of native GLP-1, native glucagon, and native GIP, but is not particularly limited thereto.
특별히 이에 제한되는 것은 아니나, 이러한 상기 펩타이드는 비자연적으로 발생된 (non-naturally occurring) 것일 수 있다.Although not particularly limited thereto, the peptide may be non-naturally occurring.
이러한 펩타이드는 분자 내 가교 (intramolecular bridge)를 포함할 수 있으며 (예컨대, 공유결합적 가교 또는 비공유결합적 가교), 구체적으로 고리를 포함하는 형태일 수 있으며, 예컨대 펩타이드의 16번 및 20번 위치의 아미노산 사이에 고리가 형성된 형태일 수 있으나, 특별히 이에 제한되는 것은 아니다. 구체적인 예로, 16번 위치의 아미노산은 글루탐산, 20번 위치의 아미노산은 리신일 수 있으나, 이에 제한되지 않는다.Such a peptide may include an intramolecular bridge (eg, a covalent bridge or a non-covalent bridge), and specifically may be in a form containing a ring, for example, at positions 16 and 20 of the peptide. It may be in the form of a ring formed between amino acids, but is not particularly limited thereto. As a specific example, the amino acid at position 16 may be glutamic acid, and the amino acid at position 20 may be lysine, but is not limited thereto.
상기 고리의 비제한적인 예로 락탐 가교 (또는 락탐 고리)를 포함할 수 있다.Non-limiting examples of the ring may include a lactam bridge (or lactam ring).
또한, 상기 펩타이드는 고리를 포함하도록, 목적하는 위치에 고리를 형성할 수 있는 아미노산을 포함하도록 변형된 것을 모두 포함한다. In addition, the peptide includes all those modified to include a ring and an amino acid capable of forming a ring at a desired position.
예컨대, 펩타이드의 16번 및 20번 위치의 아미노산 쌍이 각각 고리를 형성할 수 있는 글루탐산 또는 리신으로 치환된 것일 수 있으나, 이에 제한되지 않는다. For example, the amino acid pair at positions 16 and 20 of the peptide may be substituted with glutamic acid or lysine capable of forming a ring, respectively, but is not limited thereto.
이러한 고리는 상기 펩타이드 내의 아미노산 곁 사슬 간에 형성될 수 있으며, 그 예로 리신의 곁 사슬과 글루탐산의 곁 사슬 간에 락탐 고리가 형성되는 형태일 수 있으나, 특별히 이에 제한되는 것은 아니다. Such a ring may be formed between amino acid side chains in the peptide, for example, a lactam ring may be formed between a lysine side chain and a glutamic acid side chain, but is not particularly limited thereto.
이러한 방법들의 조합으로 제조되는 펩타이드의 예로, 천연형 글루카곤과 아미노산 서열이 하나 이상 다르고, N-말단의 아미노산 잔기의 알파-탄소가 제거된, 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 보유한 펩타이드 등이 있으나, 이에 제한되지 않으며, 아날로그 제조를 위한 여러 방법들의 조합으로 본 발명에 적용되는 펩타이드를 제조할 수 있다. As an example of a peptide prepared by a combination of these methods, the amino acid sequence is different from that of native glucagon by one or more, and the alpha-carbon of the N-terminal amino acid residue has been removed, glucagon receptor, GLP-1 receptor, and activity against GIP receptor There are peptides having a
또한, 특별히 이에 제한되지 않으나, 본 발명의 펩타이드는 체내 반감기의 증가를 위해 활성체 분해 효소의 인식작용을 회피하기 위하여 일부 아미노산을 타 아미노산 혹은 비 천연형 화합물로 치환할 수 있다. In addition, although not particularly limited thereto, in the peptide of the present invention, some amino acids may be substituted with other amino acids or non-natural compounds in order to avoid the recognition action of an activator degrading enzyme in order to increase the half-life in the body.
구체적으로, 상기 펩타이드의 아미노산 서열 중 두 번째 아미노산 서열의 치환을 통해 분해효소의 인식 작용을 회피하여 체내 반감기를 증가시킨 펩타이드일 수 있으나, 체내 분해 효소의 인식 작용을 회피하기 위한 아미노산 치환 또는 변경은 제한 없이 포함된다.Specifically, the peptide may be a peptide having an increased half-life in the body by avoiding the recognition action of the degrading enzyme through substitution of the second amino acid sequence among the amino acid sequence of the peptide, but amino acid substitution or change to avoid the recognition action of the degrading enzyme in the body included without limitation.
또한, 펩타이드 제조를 위한 이러한 변형은 L-형 혹은 D-형 아미노산, 및/또는 비-천연형 아미노산을 이용한 변형; 및/또는 천연형 서열을 개질, 예를 들어 측쇄 작용기의 변형, 분자 내 공유결합, 예컨대, 측쇄 간 고리 형성, 메틸화, 아실화, 유비퀴틴화, 인산화, 아미노헥산화, 바이오틴화 등과 같이 개질함으로써 변형하는 것을 모두 포함한다. In addition, such modifications for the production of peptides include modifications with L- or D-form amino acids, and/or non-natural amino acids; and/or by modifying the native sequence, e.g., modification of side chain functional groups, intramolecular covalent bonds, such as inter-side chain ring formation, methylation, acylation, ubiquitination, phosphorylation, aminohexylation, biotinylation, etc. includes all that
또한, 천연형 글루카곤의 아미노 및/또는 카르복시 말단에 하나 또는 그 이상의 아미노산이 추가된 것을 모두 포함한다. In addition, it includes all those in which one or more amino acids are added to the amino and/or carboxy terminus of native glucagon.
상기 치환되거나 추가되는 아미노산은 인간 단백질에서 통상적으로 관찰되는 20개의 아미노산뿐만 아니라 비정형 또는 비-자연적 발생 아미노산을 사용할 수 있다. 비정형 아미노산의 상업적 출처에는 Sigma-Aldrich, ChemPep과 Genzyme pharmaceuticals가 포함된다. 이러한 아미노산이 포함된 펩타이드와 정형적인 펩타이드 서열은 상업화된 펩타이드 합성 회사, 예를 들어 미국의 American peptide company나 Bachem, 또는 한국의 Anygen을 통해 합성 및 구매 가능하다. The amino acids to be substituted or added may be atypical or non-naturally occurring amino acids as well as the 20 amino acids commonly observed in human proteins. Commercial sources of atypical amino acids include Sigma-Aldrich, ChemPep and Genzyme Pharmaceuticals. Peptides containing these amino acids and canonical peptide sequences can be synthesized and purchased from commercial peptide synthesis companies, for example, American peptide company or Bachem in the United States, or Anygen in Korea.
아미노산 유도체도 마찬가지 방식으로 입수할 수 있는데, 그 예를 일부만 들자면 4-이미다조아세트산 (4-imidazoacetic acid) 등을 사용할 수 있다. Amino acid derivatives can also be obtained in the same manner, and 4-imidazoacetic acid can be used, to name just a few examples.
또한, 본 발명에 따른 펩타이드는 생체 내의 단백질 절단 효소들로부터 보호하고 안정성을 증가시키기 위하여 이의 N-말단 및/또는 C-말단 등이 화학적으로 수식되거나 유기단으로 보호되거나, 또는 펩타이드 말단 등에 아미노산이 추가되어 변형된 형태일 수 있다. In addition, the peptide according to the present invention has its N-terminus and/or C-terminus chemically modified or protected by an organic group in order to protect it from proteolytic enzymes in vivo and to increase stability, or amino acids are added to the peptide terminus, etc. It may be added and modified form.
특히, 화학적으로 합성한 펩타이드의 경우, N- 및 C-말단이 전하를 띠고 있기 때문에, 이러한 전하를 제거하기 위하여 N-말단을 아세틸화 (acetylation) 및/또는 C-말단을 아미드화 (amidation)할 수 있으나, 특별히 이에 제한되지는 않는다. In particular, in the case of a chemically synthesized peptide, since the N- and C-terminus are charged, the N-terminus is acetylated and/or the C-terminus is amidated to remove these charges. However, it is not particularly limited thereto.
또한, 본 발명에 따른 펩타이드는 펩타이드 그 자체, 이의 염 (예컨대, 상기 펩타이드의 약학적으로 허용가능한 염), 또는 이의 용매화물의 형태를 모두 포함한다. 또한, 펩타이드는 약학적으로 허용되는 임의의 형태일 수 있다. In addition, the peptide according to the present invention includes both the peptide itself, a salt thereof (eg, a pharmaceutically acceptable salt of the peptide), or a solvate thereof. In addition, the peptide may be in any pharmaceutically acceptable form.
상기 염의 종류는 특별히 제한되지는 않는다. 다만, 개체, 예컨대 포유류에게 안전하고 효과적인 형태인 것이 바람직하나, 특별히 이에 제한되는 것은 아니다. The type of the salt is not particularly limited. However, it is preferable that the form is safe and effective for an individual, such as a mammal, but is not particularly limited thereto.
상기 용어, "약학적으로 허용되는"은 의약학적 판단의 범위 내에서，과도한 독성，자극, 또는 알레르기 반응 등을 유발하지 않고 원하는 용도에 효과적으로 사용 가능한 물질을 의미한다. The term, "pharmaceutically acceptable" means a substance that can be effectively used for a desired purpose without causing excessive toxicity, irritation, or allergic reaction within the scope of medical judgment.
본 발명에서 용어, "약학적으로 허용되는 염" 이란 약학적으로 허용되는 무기산, 유기산, 또는 염기로부터 유도된 염을 포함한다. 적합한 산의 예로는 염산, 브롬산, 황산, 질산, 과염소산, 푸마르산，말레산, 인산, 글리콜산, 락트산, 살리실산，숙신산, 톨루엔-p-설폰산, 타르타르산, 아세트산，시트르산, 메탄설폰산，포름산, 벤조산, 말론산, 나프탈렌-2-설폰산，벤젠설폰산 등을 들 수 있다. 적합한 염기로부터 유도된 염은 나트륨, 칼륨 등의 알칼리 금속，마그네슘 등의 알칼리 토금속，및 암모늄 등을 포함할 수 있다.As used herein, the term "pharmaceutically acceptable salt" includes salts derived from pharmaceutically acceptable inorganic acids, organic acids, or bases. Examples of suitable acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfonic acid, tartaric acid, acetic acid, citric acid, methanesulfonic acid, formic acid , benzoic acid, malonic acid, naphthalene-2-sulfonic acid, benzenesulfonic acid, and the like. Salts derived from suitable bases may include alkali metals such as sodium and potassium, alkaline earth metals such as magnesium, and ammonium.
또한, 본 발명에서 사용된 용어 "용매화물"은 본 발명에 따른 펩타이드 또는 이의 염이 용매 분자와 복합체를 형성한 것을 말한다. Also, as used herein, the term “solvate” refers to a compound in which the peptide or salt thereof according to the present invention forms a complex with a solvent molecule.
하나의 구체예로서, 상기 펩타이드 (3중 작용제), 또는 화학식 1의 Z는 하기 일반식 1로 표시된 아미노산 서열을 포함할 수 있다. In one embodiment, the peptide (triple agonist), or Z of Formula 1 may include an amino acid sequence represented by Formula 1 below.
Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (일반식 1, 서열번호 103)Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp23 Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (Formula 1, SEQ ID NO: 103)
상기 일반식 1에서,In the above general formula 1,
Xaa1은 히스티딘, 4-이미다조아세틸, 또는 티로신이고, Xaa1 is histidine, 4-imidazoacetyl, or tyrosine;
Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이며,Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
Xaa3은 글루탐산 또는 글루타민이고, Xaa3 is glutamic acid or glutamine,
Xaa7은 트레오닌 또는 이소류신이며,Xaa7 is threonine or isoleucine,
Xaa10은 류신, 티로신, 리신, 시스테인, 또는 발린이고, Xaa10 is leucine, tyrosine, lysine, cysteine, or valine;
Xaa12는 리신, 세린, 또는 이소류신이며, Xaa12 is lysine, serine, or isoleucine,
Xaa13은 글루타민, 티로신, 알라닌, 또는 시스테인이고, Xaa13 is glutamine, tyrosine, alanine, or cysteine;
Xaa14는 류신, 메티오닌, 또는 티로신이며,Xaa14 is leucine, methionine, or tyrosine,
Xaa15는 시스테인, 아스파르트산, 글루탐산, 또는 류신이며,Xaa15 is cysteine, aspartic acid, glutamic acid, or leucine,
Xaa16은 글리신, 글루탐산, 또는 세린이고, Xaa16 is glycine, glutamic acid, or serine;
Xaa17은 글루타민, 아르기닌, 이소류신, 글루탐산, 시스테인, 또는 리신이며,Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine;
Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고,Xaa18 is alanine, glutamine, arginine, or histidine;
Xaa19는 알라닌, 글루타민, 시스테인, 또는 발린이며,Xaa19 is alanine, glutamine, cysteine, or valine;
Xaa20은 리신, 글루타민, 또는 아르기닌이고,Xaa20 is lysine, glutamine, or arginine;
Xaa21은 글루탐산, 글루타민, 류신, 시스테인, 또는 아스파르트산이며,Xaa21 is glutamic acid, glutamine, leucine, cysteine, or aspartic acid;
Xaa23은 이소류신 또는 발린이고,Xaa23 is isoleucine or valine,
Xaa24는 알라닌, 글루타민, 시스테인, 아스파라긴, 아스파르트산, 또는 글루탐산이며,Xaa24 is alanine, glutamine, cysteine, asparagine, aspartic acid, or glutamic acid,
Xaa27은 발린, 류신, 또는 리신이고,Xaa27 is valine, leucine, or lysine;
Xaa28은 시스테인, 리신, 알라닌, 아스파라긴, 또는 아스파르트산이며,Xaa28 is cysteine, lysine, alanine, asparagine, or aspartic acid;
Xaa29는 시스테인, 글리신, 글루타민, 트레오닌, 글루탐산, 또는 히스티딘이고,Xaa29 is cysteine, glycine, glutamine, threonine, glutamic acid, or histidine;
Xaa30은 시스테인, 글리신, 리신, 또는 히스티딘이거나, 부존재하며,Xaa30 is cysteine, glycine, lysine, or histidine, or is absent;
R1은 시스테인, GKKNDWKHNIT (서열번호 106), m-SSGAPPPS-n (서열번호 107), 또는 m-SSGQPPPS-n (서열번호 108)이거나, 부존재하며, R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or is absent;
여기서, here,
m은 -Cys-, -Pro-, 또는 -Gly-Pro-이고, m is -Cys-, -Pro-, or -Gly-Pro-;
n은 -Cys-, -Gly-, -Ser-, 또는 -His-Gly-이거나, 부존재함.n is -Cys-, -Gly-, -Ser-, or -His-Gly-, or absent.
상기 일반식 1은 서열번호 12일 경우 Xaa27은 예외적으로 메티오닌일 수 있다.In the case of Formula 1, SEQ ID NO: 12, Xaa27 may be exceptionally methionine.
상기 삼중 활성체 (3중 작용제) 또는 화학식 1의 Z의 예로, 서열번호: 1 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하는 것, 서열번호: 1 내지 11, 13 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하는 것, 서열번호: 1 내지 11, 13 내지 102로 이루어진 군에서 선택된 아미노산 서열로 (필수적으로) 구성된 것일 수 있으나, 이에 제한되는 것은 아니다. As an example of the triple activator (triple agonist) or Z of Formula 1, one comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102, SEQ ID NOs: 1 to 11, and selected from the group consisting of 13 to 102 Those containing an amino acid sequence, SEQ ID NOs: 1 to 11, 13 to 102 may be configured (essentially) of an amino acid sequence selected from the group consisting of, but is not limited thereto.
또 하나의 구체예로, 상기 삼중 활성체 (3중 작용제) 또는 화학식 1의 Z는 서열번호 21, 22, 42, 43, 50, 64, 66, 67, 70, 71, 76, 77, 96, 97 및 100으로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, (필수적으로) 구성된 것일 수 있으나, 이에 제한되는 것은 아니다.In another embodiment, the triple active agent (triple agent) or Z of Formula 1 is SEQ ID NO: 21, 22, 42, 43, 50, 64, 66, 67, 70, 71, 76, 77, 96, It may include or (essentially) an amino acid sequence selected from the group consisting of 97 and 100, but is not limited thereto.
또 하나의 구체예로, 상기 삼중 활성체 (3중 작용제) 또는 화학식 1의 Z는 서열번호 21, 22, 42, 43, 50, 66, 67, 77, 96, 97 및 100으로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, (필수적으로) 구성된 것일 수 있으나, 이에 제한되는 것은 아니다.In another embodiment, the triple active agent (triple agent) or Z of Formula 1 is selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 66, 67, 77, 96, 97 and 100 It may include or (essentially) consist of an amino acid sequence, but is not limited thereto.
또 하나의 구체예로, 상기 삼중 활성체 (3중 작용제) 또는 화학식 1의 Z는 서열번호 21, 22, 42, 43, 50, 77 및 96으로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, (필수적으로) 구성된 것일 수 있으나, 이에 제한되는 것은 아니다.In another embodiment, the triple activator (triple agonist) or Z of Formula 1 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 77 and 96, or (essential ), but is not limited thereto.
또한, 본원에서 특정 서열번호로 '구성되는' 펩타이드라고 기재되어 있다 하더라도, 해당 서열번호의 아미노산 서열로 이루어진 펩타이드와 동일 혹은 상응하는 활성을 가지는 경우라면 해당 서열번호의 아미노산 서열 앞뒤의 무의미한 서열 추가 또는 자연적으로 발생할 수 있는 돌연변이, 혹은 이의 잠재성 돌연변이 (silent mutation)를 제외하는 것이 아니며, 이러한 서열 추가 혹은 돌연변이를 가지는 경우에도 본원의 범위 내에 속하는 것이 자명하다. 즉, 일부 서열의 차이가 있더라도 일정 수준 이상의 상동성을 나타내며 글루카곤 수용체, GLP-1 수용체, 및/또는 GIP 수용체에 대한 활성을 나타낸다면 본 발명의 범위에 속할 수 있다.In addition, even if it is described as a peptide 'consisting of' a specific SEQ ID NO in the present application, if it has the same or corresponding activity as the peptide consisting of the amino acid sequence of the SEQ ID NO: Addition of meaningless sequences before and after the amino acid sequence of the corresponding SEQ ID NO: Naturally occurring mutations or latent mutations thereof are not excluded, and it is apparent that such sequence additions or mutations fall within the scope of the present application. That is, even if there is a difference in some sequences, it may fall within the scope of the present invention if it exhibits homology at a certain level or more and exhibits activity on a glucagon receptor, a GLP-1 receptor, and/or a GIP receptor.
예를 들어, 본 발명의 펩타이드에 WO2017-116204 및 WO2017-116205를 참조할 수 있다.See, for example, WO2017-116204 and WO2017-116205 for the peptides of the present invention.
이상의 내용은 본 발명의 다른 구체예 혹은 다른 양태에도 적용될 수 있으나, 이에 제한되는 것은 아니다. The above content may be applied to other embodiments or other aspects of the present invention, but is not limited thereto.
구체적으로, 상기 일반식 1에서 Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인, 아스파르트산, 또는 류신일 수 있다. Specifically, in Formula 1, Xaa14 may be leucine or methionine, and Xaa15 may be cysteine, aspartic acid, or leucine.
이러한 펩타이드의 예로, 서열번호: 1 내지 11, 14 내지 17, 및 21 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, 이로 (필수적으로) 구성된 펩타이드를 들 수 있으나, 특별히 이에 제한되는 것은 아니다. Examples of such a peptide include, but are not particularly limited to, a peptide comprising or (essentially) consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 11, 14 to 17, and 21 to 102.
이러한 펩타이드는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체 중 하나 이상을 유의하게 활성화시킬 수 있으나, 특별히 이에 제한되는 것은 아니다. 구체적으로, GLP-1을 유의하게 활성화시키거나, 추가로 글루카곤 수용체 및/또는 GIP 수용체를 유의하게 활성화시키는 것일 수 있으나, 특별히 이에 제한되지 않는다. Such a peptide may significantly activate one or more of a glucagon receptor, a GLP-1 receptor, and a GIP receptor, but is not particularly limited thereto. Specifically, it may significantly activate GLP-1 or further significantly activate glucagon receptor and/or GIP receptor, but is not particularly limited thereto.
하나의 구체예로, In one embodiment,
상기 일반식 1에서,In the above general formula 1,
Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이고,Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
Xaa7은 트레오닌이며,Xaa7 is threonine,
Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa10 is tyrosine, cysteine, or valine,
Xaa12는 리신 또는 이소류신이며, Xaa12 is lysine or isoleucine,
Xaa13은 티로신, 알라닌, 글루타민, 또는 시스테인이고, Xaa13 is tyrosine, alanine, glutamine, or cysteine;
Xaa14는 류신, 티로신, 또는 메티오닌이며, Xaa14 is leucine, tyrosine, or methionine;
Xaa15는 시스테인, 류신, 글루탐산, 또는 아스파르트산이고,Xaa15 is cysteine, leucine, glutamic acid, or aspartic acid;
Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 글루탐산, 또는 리신이며,Xaa17 is glutamine, arginine, isoleucine, cysteine, glutamic acid, or lysine;
Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고,Xaa18 is alanine, glutamine, arginine, or histidine;
Xaa19는 알라닌, 글루타민, 발린, 또는 시스테인이며,Xaa19 is alanine, glutamine, valine, or cysteine,
Xaa20은 리신, 아르기닌, 또는 글루타민이고,Xaa20 is lysine, arginine, or glutamine;
Xaa21은 글루탐산, 글루타민, 류신, 시스테인, 또는 아스파르트산이며,Xaa21 is glutamic acid, glutamine, leucine, cysteine, or aspartic acid;
Xaa23은 이소류신 또는 발린이고,Xaa23 is isoleucine or valine,
Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 글루탐산, 또는 아스파르트산이며,Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid, or aspartic acid;
Xaa27은 류신 또는 리신인, 펩타이드일 수 있으나, 특별히 이에 제한되는 것은 아니다. Xaa27 may be a peptide, which is leucine or lysine, but is not particularly limited thereto.
또 하나의 구체예로, In another embodiment,
상기 일반식 1에서,In the above general formula 1,
Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이고,Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
Xaa7은 트레오닌이며,Xaa7 is threonine,
Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa10 is tyrosine, cysteine, or valine,
Xaa12는 리신 또는 이소류신이며, Xaa12 is lysine or isoleucine,
Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa13 is tyrosine, alanine, or cysteine,
Xaa14는 류신 또는 메티오닌이며,Xaa14 is leucine or methionine,
Xaa15는 시스테인 또는 아스파르트산이고,Xaa15 is cysteine or aspartic acid,
Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 또는 리신이며,Xaa17 is glutamine, arginine, isoleucine, cysteine, or lysine;
Xaa18은 알라닌, 아르기닌, 또는 히스티딘이고,Xaa18 is alanine, arginine, or histidine;
Xaa19는 알라닌, 글루타민, 또는 시스테인이며,Xaa19 is alanine, glutamine, or cysteine,
Xaa20은 리신 또는 글루타민이고,Xaa20 is lysine or glutamine,
Xaa21은 글루탐산, 시스테인, 또는 아스파르트산이며,Xaa21 is glutamic acid, cysteine, or aspartic acid;
Xaa23은 발린이고,Xaa23 is valine,
Xaa24는 알라닌, 글루타민, 시스테인, 아스파라긴, 또는 아스파르트산이며,Xaa24 is alanine, glutamine, cysteine, asparagine, or aspartic acid;
Xaa27은 류신 또는 리신일 수 있으나, 특별히 이에 제한되지는 않는다. Xaa27 may be leucine or lysine, but is not particularly limited thereto.
또 하나의 구체예로, In another embodiment,
상기 일반식 1에서,In the above general formula 1,
Xaa2는 알파-메틸-글루탐산 또는 Aib이고,Xaa2 is alpha-methyl-glutamic acid or Aib;
Xaa7은 트레오닌이며,Xaa7 is threonine,
Xaa10은 티로신 또는 시스테인이고,Xaa10 is tyrosine or cysteine,
Xaa12는 리신 또는 이소류신이며, Xaa12 is lysine or isoleucine,
Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa13 is tyrosine, alanine, or cysteine,
Xaa14는 류신 또는 메티오닌이며,Xaa14 is leucine or methionine,
Xaa15는 시스테인 또는 아스파르트산이고,Xaa15 is cysteine or aspartic acid,
Xaa16은 글루탐산이며,Xaa16 is glutamic acid,
Xaa17은 아르기닌, 이소류신, 시스테인, 또는 리신이고,Xaa17 is arginine, isoleucine, cysteine, or lysine;
Xaa18은 알라닌, 아르기닌, 또는 히스티딘이며,Xaa18 is alanine, arginine, or histidine;
Xaa19는 알라닌, 글루타민, 또는 시스테인이고,Xaa19 is alanine, glutamine, or cysteine;
Xaa20은 리신 또는 글루타민이며,Xaa20 is lysine or glutamine,
Xaa21은 글루탐산 또는 아스파르트산이고,Xaa21 is glutamic acid or aspartic acid,
Xaa23은 발린이며,Xaa23 is valine,
Xaa24는 글루타민, 아스파라긴, 또는 아스파르트산이고,Xaa24 is glutamine, asparagine, or aspartic acid;
Xaa27은 류신이며,Xaa27 is leucine,
Xaa28은 시스테인, 알라닌, 아스파라긴, 또는 아스파르트산일 수 있다. Xaa28 may be cysteine, alanine, asparagine, or aspartic acid.
또 하나의 구체예로, In another embodiment,
상기 일반식 1에서, In the above general formula 1,
Xaa1은 히스티딘 또는 4-이미다조아세틸이고, Xaa1 is histidine or 4-imidazoacetyl;
Xaa2는 알파-메틸-글루탐산 또는 Aib이며,Xaa2 is alpha-methyl-glutamic acid or Aib;
Xaa3은 글루타민이고, Xaa3 is glutamine,
Xaa7은 트레오닌이며,Xaa7 is threonine,
Xaa10은 티로신이고, Xaa10 is tyrosine,
Xaa12는 이소류신이며, Xaa12 is isoleucine,
Xaa13은 알라닌 또는 시스테인이고, Xaa13 is alanine or cysteine,
Xaa14는 메티오닌이며,Xaa14 is methionine,
Xaa15는 아스파르트산이고,Xaa15 is aspartic acid,
Xaa16은 글루탐산이며,Xaa16 is glutamic acid,
Xaa17은 이소류신 또는 리신이고,Xaa17 is isoleucine or lysine,
Xaa18은 알라닌 또는 히스티딘이며,Xaa18 is alanine or histidine,
Xaa19는 글루타민 또는 시스테인이고,Xaa19 is glutamine or cysteine,
Xaa20은 리신이며,Xaa20 is lysine,
Xaa21은 아스파르트산이고,Xaa21 is aspartic acid,
Xaa23은 발린이며,Xaa23 is valine,
Xaa24는 아스파라긴이고,Xaa24 is asparagine,
Xaa27은 류신이며,Xaa27 is leucine,
Xaa28은 알라닌 또는 아스파라긴이고,Xaa28 is alanine or asparagine,
Xaa29는 글루타민 또는 트레오닌이며,Xaa29 is glutamine or threonine,
Xaa30은 시스테인 또는 리신이거나, 부존재할 수 있다. Xaa30 may be cysteine or lysine or absent.
또 하나의 구체예로,In another embodiment,
상기 일반식 1에서, In the above general formula 1,
Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이며,Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
Xaa3은 글루타민이고, Xaa3 is glutamine,
Xaa7은 트레오닌이며,Xaa7 is threonine,
Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa10 is tyrosine, cysteine, or valine,
Xaa12는 리신이며, Xaa12 is lysine,
Xaa13은 티로신이고, Xaa13 is tyrosine,
Xaa14는 류신이며,Xaa14 is leucine,
Xaa15는 아스파르트산이며,Xaa15 is aspartic acid,
Xaa16은 글리신, 글루탐산, 또는 세린이고, Xaa16 is glycine, glutamic acid, or serine;
Xaa17은 글루타민, 아르기닌, 시스테인, 또는 리신이며,Xaa17 is glutamine, arginine, cysteine, or lysine,
Xaa18은 알라닌, 아르기닌, 또는 히스티딘이고,Xaa18 is alanine, arginine, or histidine;
Xaa19는 알라닌 또는 글루타민이며,Xaa19 is alanine or glutamine,
Xaa20은 리신 또는 글루타민이고,Xaa20 is lysine or glutamine,
Xaa21은 글루탐산, 시스테인, 또는 아스파르트산이며,Xaa21 is glutamic acid, cysteine, or aspartic acid;
Xaa23은 발린이고,Xaa23 is valine,
Xaa24는 알라닌, 글루타민, 또는 시스테인이며,Xaa24 is alanine, glutamine, or cysteine,
Xaa27은 류신 또는 리신이고,Xaa27 is leucine or lysine,
Xaa29는 글리신, 글루타민, 트레오닌, 또는 히스티딘일 수 있으나, 특별히 이에 제한되는 것은 아니다. Xaa29 may be glycine, glutamine, threonine, or histidine, but is not particularly limited thereto.
이러한 펩타이드는 GLP-1 수용체 및 글루카곤 수용체의 활성화 정도가 유의하고, GIP 수용체의 활성화 정도에 비해 높거나; GLP-1 수용체, 글루카곤 수용체 및 GIP 수용체의 활성화 정도가 모두 유의하거나; GLP-1 수용체 및 GIP 수용체의 활성화 정도가 유의하고, 글루카곤 수용체의 활성화에 비해 높은 경우에 해당할 수 있으나, 특별히 이에 제한되지 않는다. These peptides have significant activation levels of GLP-1 receptors and glucagon receptors, and are higher than those of GIP receptors; The activation levels of the GLP-1 receptor, the glucagon receptor and the GIP receptor are all significant; The degree of activation of the GLP-1 receptor and the GIP receptor is significant and may correspond to a case where the activation level of the glucagon receptor is higher than that of the glucagon receptor, but is not particularly limited thereto.
이러한 펩타이드의 예로, 서열번호: 8, 9, 21 내지 37, 39, 42, 43, 49 내지 61, 64 내지 83, 85, 86, 88, 89, 91 내지 93, 95 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, 이로 (필수적으로) 구성된 펩타이드를 들 수 있으나, 특별히 이에 제한되는 것은 아니다. Examples of such peptides include SEQ ID NOs: 8, 9, 21 to 37, 39, 42, 43, 49 to 61, 64 to 83, 85, 86, 88, 89, 91 to 93, selected from the group consisting of 95 to 102 and a peptide comprising or (essentially) consisting of an amino acid sequence, but is not particularly limited thereto.
구체적인 양태로서, 상기 펩타이드는 하기 일반식 2로 표시되는 아미노산 서열을 포함하는 것일 수 있다. In a specific embodiment, the peptide may include an amino acid sequence represented by the following general formula (2).
Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (일반식 2, 서열번호 104)Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (General Formula 2, SEQ ID NO: 104)
상기 식에서,In the above formula,
Xaa1은 4-이미다조아세틸, 히스티딘, 또는 티로신이고;Xaa1 is 4-imidazoacetyl, histidine, or tyrosine;
Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이며;Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib;
Xaa10은 티로신, 또는 시스테인이며Xaa10 is tyrosine or cysteine
Xaa13은 알라닌, 글루타민, 티로신, 또는 시스테인이며;Xaa13 is alanine, glutamine, tyrosine, or cysteine;
Xaa14는 류신, 메티오닌, 또는 티로신이고;Xaa14 is leucine, methionine, or tyrosine;
Xaa15는 아스파르트산, 글루탐산, 또는 류신이며;Xaa15 is aspartic acid, glutamic acid, or leucine;
Xaa16은 글리신, 글루탐산, 또는 세린이고;Xaa16 is glycine, glutamic acid, or serine;
Xaa17은 글루타민, 아르기닌, 이소류신, 글루탐산, 시스테인, 또는 리신이며;Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine;
Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고;Xaa18 is alanine, glutamine, arginine, or histidine;
Xaa19는 알라닌, 글루타민, 시스테인, 또는 발린이며;Xaa19 is alanine, glutamine, cysteine, or valine;
Xaa20은 리신, 글루타민, 또는 아르기닌이고;Xaa20 is lysine, glutamine, or arginine;
Xaa21은 시스테인, 글루탐산, 글루타민, 류신, 또는 아스파르트산이며;Xaa21 is cysteine, glutamic acid, glutamine, leucine, or aspartic acid;
Xaa23은 이소류신 또는 발린이고;Xaa23 is isoleucine or valine;
Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 또는 글루탐산이며;Xaa24 is cysteine, alanine, glutamine, asparagine, or glutamic acid;
Xaa28은 리신, 시스테인, 아스파라긴, 또는 아스파르트산이며;Xaa28 is lysine, cysteine, asparagine, or aspartic acid;
Xaa29는 글리신, 글루타민, 시스테인, 또는 히스티딘이고;Xaa29 is glycine, glutamine, cysteine, or histidine;
Xaa30은 시스테인, 글리신, 리신, 또는 히스티딘이며;Xaa30 is cysteine, glycine, lysine, or histidine;
Xaa31은 프롤린 또는 시스테인이며;Xaa31 is proline or cysteine;
Xaa40은 시스테인이거나, 부존재함.Xaa40 is cysteine or absent.
보다 구체적으로, 상기 일반식 2에서,More specifically, in the general formula 2,
Xaa13은 알라닌, 티로신, 또는 시스테인이며;Xaa13 is alanine, tyrosine, or cysteine;
Xaa15는 아스파르트산 또는 글루탐산이고,Xaa15 is aspartic acid or glutamic acid,
Xaa17은 글루타민, 아르기닌, 시스테인, 또는 리신이며;Xaa17 is glutamine, arginine, cysteine, or lysine;
Xaa18은 알라닌, 아르기닌, 또는 히스티딘이고;Xaa18 is alanine, arginine, or histidine;
Xaa21은 시스테인, 글루탐산, 글루타민, 또는 아스파르트산이며;Xaa21 is cysteine, glutamic acid, glutamine, or aspartic acid;
Xaa23은 이소류신 또는 발린이고;Xaa23 is isoleucine or valine;
Xaa24는 시스테인, 글루타민, 또는 아스파라긴이고, Xaa24 is cysteine, glutamine, or asparagine;
Xaa28은 시스테인, 아스파라긴, 또는 아스파르트산이며;Xaa28 is cysteine, asparagine, or aspartic acid;
Xaa29는 글루타민, 시스테인, 또는 히스티딘이고;Xaa29 is glutamine, cysteine, or histidine;
Xaa30은 시스테인, 리신, 또는 히스티딘일 수 있다. Xaa30 may be cysteine, lysine, or histidine.
이러한 펩타이드의 예로, 서열번호: 21, 22, 42, 43, 50, 64 내지 77, 및 95 내지 102로 이루어진 군에서 선택된 아미노산 서열, 또는 서열번호: 21, 22, 42, 43, 50, 64 내지 77, 및 96 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, 이로 (필수적으로) 구성된 펩타이드를 들 수 있으나, 특별히 이에 제한되는 것은 아니다. Examples of such a peptide include an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 64 to 77, and 95 to 102, or SEQ ID NOs: 21, 22, 42, 43, 50, 64 to 77, and a peptide comprising or (essentially) consisting of an amino acid sequence selected from the group consisting of 96 to 102, but is not particularly limited thereto.
구체적인 양태로서, 상기 펩타이드는 하기 일반식 3의 아미노산 서열을 포함할 수 있다. In a specific embodiment, the peptide may include an amino acid sequence of the following general formula (3).
Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (일반식 3, 서열번호 105),Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp- Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (General Formula 3, SEQ ID NO: 105);
상기 일반식 3에서,In the above general formula 3,
Xaa1은 히스티딘 또는 티로신이고;Xaa1 is histidine or tyrosine;
Xaa2는 알파-메틸-글루탐산 또는 Aib이며;Xaa2 is alpha-methyl-glutamic acid or Aib;
Xaa13은 알라닌, 티로신 또는 시스테인이고;Xaa13 is alanine, tyrosine or cysteine;
Xaa17은 아르기닌, 시스테인, 또는 리신이며;Xaa17 is arginine, cysteine, or lysine;
Xaa18은 알라닌 또는 아르기닌이고;Xaa18 is alanine or arginine;
Xaa19는 알라닌 또는 시스테인이며;Xaa19 is alanine or cysteine;
Xaa21은 글루탐산 또는 아스파르트산이고;Xaa21 is glutamic acid or aspartic acid;
Xaa24는 글루타민 또는 아스파라긴이며,Xaa24 is glutamine or asparagine,
Xaa28은 시스테인 또는 아스파르트산이며;Xaa28 is cysteine or aspartic acid;
Xaa29는 시스테인, 히스티딘, 또는 글루타민이고;Xaa29 is cysteine, histidine, or glutamine;
Xaa30은 시스테인 또는 히스티딘이며;Xaa30 is cysteine or histidine;
Xaa31은 프롤린 또는 시스테인이며;Xaa31 is proline or cysteine;
Xaa40은 시스테인 또는 부존재할 수 있다. Xaa40 may be cysteine or absent.
이러한 펩타이드의 예로, 서열번호: 21, 22, 42, 43, 50, 64 내지 71, 75 내지 77, 및 96 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, 이로 (필수적으로) 구성된 펩타이드를 들 수 있으나, 특별히 이에 제한되는 것은 아니다. Examples of such peptides include peptides comprising or (essentially) consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 64 to 71, 75 to 77, and 96 to 102. However, it is not particularly limited thereto.
또한, 상기 일반식 1에서 R1은 시스테인, GKKNDWKHNIT(서열번호 106), CSSGQPPPS (서열번호 109), GPSSGAPPPS (서열번호 110), GPSSGAPPPSC (서열번호 111), PSSGAPPPS (서열번호 112), PSSGAPPPSG (서열번호 113), PSSGAPPPSHG (서열번호 114), PSSGAPPPSS (서열번호 115), PSSGQPPPS (서열번호 116), 또는 PSSGQPPPSC (서열번호 117)이거나, 부존재할 수 있으나, 특별히 이에 제한되는 것은 아니다. In addition, in Formula 1, R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), CSSGQPPPS (SEQ ID NO: 109), GPSSGAPPPS (SEQ ID NO: 110), GPSSGAPPPSC (SEQ ID NO: 111), PSSGAPPPS (SEQ ID NO: 112), PSSGAPPPSG (SEQ ID NO: SEQ ID NO: 110) 113), PSSGAPPPSHG (SEQ ID NO: 114), PSSGAPPPSS (SEQ ID NO: 115), PSSGQPPPS (SEQ ID NO: 116), or PSSGQPPPSC (SEQ ID NO: 117), or absent, but is not particularly limited thereto.
또한, 본 발명의 펩타이드는 그 길이에 따라 이 분야에서 잘 알려진 방법, 예를 들어 자동 펩타이드 합성기에 의해 합성할 수 있으며, 유전자 조작 기술에 의하여 생산할 수도 있다. In addition, according to the length of the peptide of the present invention, it can be synthesized by a method well known in the art, for example, an automatic peptide synthesizer, or it can be produced by a genetic engineering technique.
구체적으로, 본 발명의 펩타이드는 표준 합성 방법, 재조합 발현 시스템, 또는 임의의 다른 당해 분야의 방법에 의해 제조될 수 있다. 따라서, 본 발명에 따른 펩타이드는, 예를 들어 하기를 포함하는 방법을 포함하는 다수의 방법으로 합성될 수 있다:Specifically, the peptides of the present invention can be prepared by standard synthetic methods, recombinant expression systems, or any other method in the art. Thus, the peptides according to the invention can be synthesized in a number of ways, including, for example, those comprising:
(a) 펩타이드를 고체상 또는 액체상 방법의 수단으로 단계적으로 또는 단편 조립에 의해 합성하고, 최종 펩타이드 생성물을 분리 및 정제하는 방법; 또는(a) a method for synthesizing peptides stepwise or by fragment assembly by means of solid or liquid phase methods, and isolating and purifying the final peptide product; or
(b) 펩타이드를 인코딩하는 핵산 작제물을 숙주세포 내에서 발현시키고, 발현 생성물을 숙주 세포 배양물로부터 회수하는 방법; 또는(b) expressing a nucleic acid construct encoding the peptide in a host cell and recovering the expression product from the host cell culture; or
(c) 펩타이드를 인코딩하는 핵산 작제물의 무세포 시험관 내 발현을 수행하고, 발현 생성물을 회수하는 방법; 또는 (c) performing cell-free in vitro expression of a nucleic acid construct encoding the peptide and recovering the expression product; or
(a), (b) 및 (c)의 임의의 조합으로 펩타이드의 단편을 수득하고, 이어서 단편을 연결시켜 펩타이드를 수득하고, 당해 펩타이드를 회수하는 방법.A method for obtaining a fragment of a peptide by any combination of (a), (b) and (c), and then ligating the fragments to obtain a peptide, and recovering the peptide.
한편, 3중 작용성 지속형 결합체 또는 3중 작용제는 비자연적인 (non-naturally occurring) 것일 수 있다. Meanwhile, the triple-acting long-acting binder or the triple-acting agent may be non-naturally occurring.
본 발명의 하나의 구체예에서, 상기 3중 작용성 지속형 결합체는 상기에서 기재한 화학식 1로 표시되는, 결합체일 수 있으나, 이에 제한되지 않는다.In one embodiment of the present invention, the triple-functional long-acting conjugate may be a conjugate represented by Formula 1 described above, but is not limited thereto.
구체적으로, 화학식 1을 설명하면 다음과 같다.Specifically, Formula 1 will be described as follows.
[화학식 1][Formula 1]
Z - Lx - FcZ - Lx - Fc
이 때 Lx는 에틸렌글리콜 반복 단위를 함유하는 링커이며, x는 0 또는 자연수이고,In this case, Lx is a linker containing an ethylene glycol repeating unit, and x is 0 or a natural number,
Fc는 면역글로불린 Fc 영역이고, Fc is an immunoglobulin Fc region,
-는 Lx와 Z 사이, Fc와 Lx 사이의 각각에 공유결합 연결을 나타내며, - represents a covalent linkage between Lx and Z, between Fc and Lx, respectively,
Z는 상기에서 설명한 바와 같다. Z는 3중 작용제를 의미할 수 있다.Z is as described above. Z may mean triple agonist.
상기 결합체에서 Fc는 Z, 즉 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는, 펩타이드의 반감기를 증가시킬 수 있는 물질로서, 본 발명의 상기 결합체를 구성하는 모이어티의 일 구성에 해당한다. In the conjugate, Fc is a substance capable of increasing the half-life of a peptide, having activity against Z, that is, glucagon receptor, GLP-1 receptor, and GIP receptor, in one component of the moiety constituting the conjugate of the present invention. corresponds to
상기 Fc는 Z와 공유 화학결합 또는 비공유 화학결합으로 서로 결합되는 것일 수 있으며, 공유 화학결합, 비공유 화학결합 또는 이들의 조합으로 Lx를 통하여 Fc와 Z가 서로 결합되는 것일 수 있다.The Fc may be bonded to each other by a covalent chemical bond or a non-covalent chemical bond with Z, and Fc and Z may be bonded to each other through Lx by a covalent chemical bond, a non-covalent chemical bond, or a combination thereof.
상기 Fc는 Z와 직접적으로 연결되거나 (즉, 상기 화학식 1에서 x가 0이거나), 또는 링커 (Lx)를 통해 연결된 것일 수 있다. The Fc may be directly linked to Z (ie, x is 0 in Formula 1) or linked through a linker (Lx).
구체적으로, 상기 Lx는 비펩타이드성 링커, 예를 들어 에틸렌글리콜 반복 단위를 함유하는 링커일 수 있다. Specifically, Lx may be a non-peptidyl linker, for example, a linker containing an ethylene glycol repeating unit.
본 발명에서 "비펩타이드성 링커"는 반복 단위가 2개 이상 결합된 생체 적합성 중합체를 포함한다. 상기 반복 단위들은 펩타이드 결합이 아닌 임의의 공유결합을 통해 서로 연결된다. 상기 비펩타이드성 링커는 본 발명의 결합체의 모이어티를 이루는 일 구성일 수 있으며, 상기 화학식 1에서 Lx에 해당된다. In the present invention, "non-peptidyl linker" includes a biocompatible polymer in which two or more repeating units are bonded. The repeating units are linked to each other through any covalent bond other than a peptide bond. The non-peptidyl linker may be one component constituting a moiety of the conjugate of the present invention, and corresponds to Lx in Formula 1 above.
상기 Lx에서 x는 1 이상일 수 있으며, x가 2 이상일 때 각각의 L은 독립적일 수 있다. In Lx, x may be 1 or more, and when x is 2 or more, each L may be independent.
본 발명에서 사용될 수 있는 비펩타이드성 링커는 생체 내 단백질 분해 효소에 저항성 있는 중합체이면 제한 없이 사용될 수 있다. 본 발명에서 상기 비펩타이드성 링커는 비펩타이드성 중합체와 혼용되어 사용될 수 있다.The non-peptidyl linker that can be used in the present invention may be used without limitation as long as it is a polymer resistant to proteolytic enzymes in vivo. In the present invention, the non-peptidyl linker may be used in combination with a non-peptidyl polymer.
특별히 이에 제한되지 않으나, 상기 비펩타이드성 링커는 에틸렌글리콜 반복 단위를 함유하는 링커, 예를 들어, 폴리에틸렌 글리콜일 수 있고, 또한, 당해 분야에 이미 알려진 이들의 유도체 및 당해 분야의 기술 수준에서 용이하게 제조할 수 있는 유도체들도 본 발명의 범위에 포함된다.Although not particularly limited thereto, the non-peptidyl linker may be a linker containing an ethylene glycol repeating unit, for example, polyethylene glycol, and also derivatives thereof known in the art and easily at the level of skill in the art. Derivatives that can be prepared are also included in the scope of the present invention.
상기 비펩타이드성 링커의 반복 단위는 에틸렌글리콜 반복 단위일 수 있고, 구체적으로, 상기 비펩타이드성 링커는 에틸렌글리콜 반복 단위를 포함하면서, 결합체의 제조에 이용되는 작용기를 말단에 포함하는 것일 수 있다. 본 발명에 따른 지속형 결합체는 상기 작용기를 통해 Z와 Fc가 연결된 형태일 수 있으나 이에 제한되지 않는다. 본 발명에서, 상기 비펩타이드성 링커는 2개, 또는 3개 이상의 작용기를 포함할 수 있고, 각 작용기는 동일하거나, 서로 상이할 수 있으나, 이에 제한되지 않는다.The repeating unit of the non-peptidyl linker may be an ethylene glycol repeating unit, and specifically, the non-peptidyl linker may include an ethylene glycol repeating unit and a functional group used in the preparation of the conjugate at the terminal thereof. The long-acting conjugate according to the present invention may be in a form in which Z and Fc are linked through the functional group, but is not limited thereto. In the present invention, the non-peptidyl linker may include two, or three or more functional groups, and each functional group may be the same or different from each other, but is not limited thereto.
구체적으로, 상기 링커는 하기 화학식 2로 표시되는 폴리에틸렌글리콜(PEG)일 수 있으나, 이에 제한되는 것은 아니다:Specifically, the linker may be polyethylene glycol (PEG) represented by the following formula (2), but is not limited thereto:
[화학식 2][Formula 2]
Figure PCTKR2021009210-appb-img-000001
Figure PCTKR2021009210-appb-img-000001
여기서, n= 10 내지 2400, n= 10 내지 480, 또는 n = 50 내지 250이나, 이에 제한되지 않는다. Here, n=10 to 2400, n=10 to 480, or n=50 to 250, but is not limited thereto.
상기 지속형 결합체에서 PEG 모이어티는, -(CH2CH2O)n-구조 뿐만 아니라 연결 요소와 이 -(CH2CH2O)n- 사이에 개재하는 산소 원자도 포함할 수 있으나, 이에 제한되는 것은 아니다.The PEG moiety in the long-acting conjugate may include, but is not limited to, the -(CH2CH2O)n- structure as well as an oxygen atom intervening between the linking element and the -(CH2CH2O)n-.
또한, 하나의 구체적인 실시 형태에서 상기 결합체는 일반식 1의 아미노산 서열 또는 서열번호 1 내지 102 중 어느 하나의 아미노산 서열을 포함하는 펩타이드(Z)와 면역글로불린 Fc 영역(Fc)이 에틸렌글리콜 반복 단위를 함유하는 링커를 통해 공유 결합으로 연결된 구조 일 수 있으나, 이에 제한되는 것은 아니다.In addition, in one specific embodiment, the conjugate comprises a peptide (Z) comprising the amino acid sequence of Formula 1 or an amino acid sequence of any one of SEQ ID NOs: 1 to 102 and an immunoglobulin Fc region (Fc) comprising an ethylene glycol repeating unit It may be a structure connected by a covalent bond through a linker containing it, but is not limited thereto.
상기 폴리에틸렌 글리콜은, 에틸렌 글리콜 동종 중합체, PEG 공중합체, 또는 모노메틸-치환된 PEG 중합체 (mPEG)의 형태를 모두 포괄하는 용어이나, 특별히 이에 제한되는 것은 아니다.The polyethylene glycol is a term encompassing all forms of ethylene glycol homopolymer, PEG copolymer, or monomethyl-substituted PEG polymer (mPEG), but is not particularly limited thereto.
하나의 구체예로 상기 에틸렌글리콜 반복단위는 그 예로, [OCH2CH2]n로 표시될 수 있으며, n 값은 자연수로 상기 펩타이드 결합체 내의 [OCH2CH2]n 부위의 평균 분자량, 예컨대 수평균 분자량이 O 초과 내지 약 100 kDa이 되도록 정해질 수 있으나, 이에 제한되지 않는다. 또 하나의 예로, 상기 n 값은 자연수로 상기 펩타이드 결합체 내의 [OCH2CH2]n 부위의 평균 분자량, 예컨대 수평균 분자량이 약 1 내지 약 100 kDa, 약 1 내지 약 80 kDa, 약 1 내지 약 50 kDa, 약 1 내지 약 30 kDa, 약 1 내지 약 25 kDa, 약 1 내지 약 20 kDa, 약 1 내지 약 15 kDa, 약 1 내지 약 13 kDa, 약 1 내지 약 11 kDa, 약 1 내지 약 10 kDa, 약 1 내지 약 8 kDa, 약 1 내지 약 5 kDa, 약 1 내지 약 3.4 kDa, 약 3 내지 약 30 kDa, 약 3 내지 약 27 kDa, 약 3 내지 약 25 kDa, 약 3 내지 약 22 kDa, 약 3 내지 약 20 kDa, 약 3 내지 약 18 kDa, 약 3 내지 약 16 kDa, 약 3 내지 약 15 kDa, 약 3 내지 약 13 kDa, 약 3 내지 약 11 kDa, 약 3 내지 약 10 kDa, 약 3 내지 약 8 kDa, 약 3 내지 약 5 kDa, 약 3 내지 약 3.4 kDa, 약 8 내지 약 30 kDa, 약 8 내지 약 27 kDa, 약 8 내지 약 25 kDa, 약 8 내지 약 22 kDa, 약 8 내지 약 20 kDa, 약 8 내지 약 18 kDa, 약 8 내지 약 16 kDa, 약 8 내지 약 15 kDa, 약 8 내지 약 13 kDa, 약 8 내지 약 11 kDa, 약 8 내지 약 10 kDa, 약 9 내지 약 15 kDa, 약 9 내지 약 14 kDa, 약 9 내지 약 13 kDa, 약 9 내지 약 12 kDa, 약 9 내지 약 11 kDa, 약 9.5 내지 약 10.5 kDa, 또는 약 10 kDa일 수 있으나, 이에 제한되지 않는다.In one embodiment, the ethylene glycol repeating unit may be represented by, for example, [OCH 2 CH 2 ]n, and the value of n is a natural number, the average molecular weight of the [OCH 2 CH 2 ]n site in the peptide conjugate, such as the number The average molecular weight may be set to be greater than 0 to about 100 kDa, but is not limited thereto. In another example, the n value is a natural number, and the average molecular weight of the [OCH 2 CH 2 ]n site in the peptide conjugate, for example, has a number average molecular weight of about 1 to about 100 kDa, about 1 to about 80 kDa, about 1 to about 50 kDa, about 1 to about 30 kDa, about 1 to about 25 kDa, about 1 to about 20 kDa, about 1 to about 15 kDa, about 1 to about 13 kDa, about 1 to about 11 kDa, about 1 to about 10 kDa, about 1 to about 8 kDa, about 1 to about 5 kDa, about 1 to about 3.4 kDa, about 3 to about 30 kDa, about 3 to about 27 kDa, about 3 to about 25 kDa, about 3 to about 22 kDa , about 3 to about 20 kDa, about 3 to about 18 kDa, about 3 to about 16 kDa, about 3 to about 15 kDa, about 3 to about 13 kDa, about 3 to about 11 kDa, about 3 to about 10 kDa, about 3 to about 8 kDa, about 3 to about 5 kDa, about 3 to about 3.4 kDa, about 8 to about 30 kDa, about 8 to about 27 kDa, about 8 to about 25 kDa, about 8 to about 22 kDa, about 8 to about 20 kDa, about 8 to about 18 kDa, about 8 to about 16 kDa, about 8 to about 15 kDa, about 8 to about 13 kDa, about 8 to about 11 kDa, about 8 to about 10 kDa, about 9 to about 15 kDa, about 9 to about 14 kDa, about 9 to about 13 kDa, about 9 to about 12 kDa, about 9 to about 11 kDa, about 9.5 to about 10.5 kDa, or about 10 kDa doesn't happen
본 발명에서 사용될 수 있는 비펩타이드성 링커는 생체 내 단백질 분해 효소에 저항성 있는 에틸렌글리콜 반복단위를 포함하는 중합체이면 제한 없이 사용될 수 있다. 상기 비펩타이드성 중합체의 분자량은 0 초과 약 100 kDa 범위, 약 1 내지 약 100 kDa 범위, 구체적으로 약 1 내지 약 20 kDa 범위, 또는 약 1 내지 약 10 kDa 범위이나, 이에 제한되지 않는다. 또한, 상기 Fc에 해당하는 폴리펩타이드와 결합되는 본 발명의 비펩타이드성 링커는 한 종류의 중합체뿐만 아니라 상이한 종류의 중합체들의 조합이 사용될 수도 있다. The non-peptidyl linker that can be used in the present invention may be used without limitation as long as it is a polymer including an ethylene glycol repeating unit resistant to proteolytic enzymes in vivo. The molecular weight of the non-peptidyl polymer is, but is not limited to, greater than 0, in the range of about 100 kDa, in the range of about 1 to about 100 kDa, specifically in the range of about 1 to about 20 kDa, or in the range of about 1 to about 10 kDa. In addition, for the non-peptidyl linker of the present invention that is coupled to the polypeptide corresponding to the Fc, a combination of different types of polymers as well as one type of polymer may be used.
하나의 구체적인 실시 형태에서 상기 비펩타이드성 링커의 양 말단은 각각 Fc, 예컨대 면역글로불린 Fc 영역의 아민기, 티올기, 하이드록실기 및 Z의 아민기, 티올기, 아지드기, 하이드록실기에 결합할 수 있으나, 이에 제한되지 않는다.In one specific embodiment, both ends of the non-peptidyl linker are each Fc, such as an amine group, a thiol group, a hydroxyl group, and an amine group, a thiol group, an azide group, and a hydroxyl group of an immunoglobulin Fc region. It may be combined, but is not limited thereto.
구체적으로, 상기 비펩타이드성 중합체는 양쪽 말단에 각각 Fc (예컨대, 면역글로불린 Fc 영역) 및 Z와 결합될 수 있는 반응기, 구체적으로는 면역글로불린 Fc 영역의 시스테인의 티올기; N-말단, 리신, 아르기닌, 글루타민 및/또는 히스티딘에 위치한 아민기; 및/또는 C-말단에 위치한 하이드록실기와 결합되고, Z의 시스테인의 티올기; 리신, 아르기닌, 글루타민 및/또는 히스티딘의 아민기; 아지도리신의 아지드기; 및/또는 하이드록실기와 결합될 수 있는 반응기를 포함할 수 있으나, 이에 제한되지 않는다. Specifically, the non-peptidyl polymer has a reactive group capable of binding to Fc (eg, immunoglobulin Fc region) and Z, respectively, at both ends, specifically, a thiol group of a cysteine in the immunoglobulin Fc region; an amine group located at the N-terminus, lysine, arginine, glutamine and/or histidine; and/or bonded to a hydroxyl group located at the C-terminus, and a thiol group of cysteine of Z; amine groups of lysine, arginine, glutamine and/or histidine; azide group of azidolysin; and/or a reactive group capable of bonding to a hydroxyl group, but is not limited thereto.
하나의 구체적인 실시 형태에서 상기 비펩타이드성 링커의 양 말단은 각각 Fc, 예컨대 면역글로불린 Fc 영역의 아민기 또는 티올기 및 Z의 아민기 또는 티올기에 결합할 수 있다.In one specific embodiment, both ends of the non-peptidyl linker may be respectively bound to an amine or thiol group of an Fc region, such as an immunoglobulin Fc region, and an amine or thiol group of Z.
구체적으로, 상기 비펩타이드성 중합체는 양쪽 말단에 각각 Fc (예컨대, 면역글로불린 Fc 영역) 및 Z와 결합될 수 있는 반응기, 구체적으로는 Z, 혹은 Fc (예컨대, 면역글로불린 Fc 영역)의 N-말단 또는 리신에 위치한 아민기, 또는 시스테인의 티올기와 결합될 수 있는 반응기를 포함할 수 있으나, 이에 제한되지 않는다. Specifically, the non-peptidyl polymer has a reactive group capable of binding to Fc (eg, immunoglobulin Fc region) and Z, respectively, at both ends, specifically Z, or the N-terminus of Fc (eg, immunoglobulin Fc region). Or it may include a reactive group capable of bonding to an amine group located on lysine, or a thiol group of cysteine, but is not limited thereto.
또한, Fc, 예컨대 면역글로불린 Fc 영역 및 Z와 결합될 수 있는, 상기 비펩타이드성 중합체의 반응기는 알데히드기, 말레이미드기 및 석시니미드 유도체로 구성된 군으로부터 선택되는 하나 이상일 수 있으나, 이에 제한되지 않는다. In addition, the reactive group of the non-peptide polymer capable of binding to Fc, such as an immunoglobulin Fc region and Z, may be at least one selected from the group consisting of an aldehyde group, a maleimide group and a succinimide derivative, but is not limited thereto. .
상기에서, 알데히드기로 프로피온 알데히드기 또는 부틸 알데히드기를 예로서 들 수 있으나, 이에 제한되지 않는다. In the above, the aldehyde group may be exemplified by a propionaldehyde group or a butyl aldehyde group, but is not limited thereto.
상기에서, 석시니미드 유도체로는 석시니미딜 발레르에이트, 석시니미딜 메틸부타노에이트, 석시니미딜 메틸프로피온에이트, 석시니미딜 부타노에이트, 석시니미딜 프로피오네이트, N-하이드록시석시니미드, 히드록시 석시니미딜, 석시니미딜 카르복시메틸 또는 석시니미딜 카보네이트가 이용될 수 있으나, 이에 제한되지 않는다. In the above, as the succinimide derivative, succinimidyl valerate, succinimidyl methylbutanoate, succinimidyl methylpropionate, succinimidyl butanoate, succinimidyl propionate, N-hydroxysuccini Mead, hydroxy succinimidyl, succinimidyl carboxymethyl or succinimidyl carbonate may be used, but are not limited thereto.
비펩타이드성 링커는 이러한 반응기를 통하여 Z와 Fc에 연결되어 비펩타이드성 링커 연결부로 전환될 수 있으나, 특별히 이에 제한되는 것은 아니다.The non-peptide linker may be connected to Z and Fc through such a reactive group and converted into a non-peptide linker linkage, but is not particularly limited thereto.
또한, 알데히드 결합에 의한 환원성 아민화로 생성된 최종 산물은 아미드 결합으로 연결된 것보다 훨씬 안정적이다. 알데히드 반응기는 낮은 pH에서 N-말단에 선택적으로 반응하며, 높은 pH, 예를 들어 pH 9.0 조건에서는 리신 잔기와 공유결합을 형성할 수 있다. In addition, the final product resulting from reductive amination by aldehyde bonds is much more stable than those linked by amide bonds. The aldehyde reactive group selectively reacts with the N-terminus at a low pH, and can form a covalent bond with a lysine residue at a high pH, for example, pH 9.0.
또한, 상기 비펩타이드성 링커의 양쪽 말단의 반응기는 서로 동일하거나 또는 서로 상이할 수 있으며, 예를 들어, 한쪽 말단에는 말레이미드기를, 다른 쪽 말단에는 알데히드기, 프로피온 알데히드기, 또는 부틸 알데히드기를 가질 수 있다. 그러나, 비펩타이드성 링커의 각 말단에 Fc, 구체적으로 면역글로불린 Fc 영역과 Z가 결합될 수 있다면, 특별히 이에 제한되지 않는다. In addition, the reactive groups at both ends of the non-peptidyl linker may be the same or different from each other, for example, a maleimide group at one end and an aldehyde group, a propionaldehyde group, or a butyl aldehyde group at the other end. . However, if Fc, specifically, immunoglobulin Fc region and Z can be bound to each end of the non-peptidyl linker, it is not particularly limited thereto.
예를 들어, 상기 비펩타이드성 링커의 한쪽 말단에는 반응기로서 말레이미드 기를 포함하고, 다른 쪽 말단에는 알데히드기, 프로피온 알데히드기 또는 부틸 알데히드기 등을 포함할 수 있다. For example, one end of the non-peptidyl linker may include a maleimide group as a reactive group, and an aldehyde group, a propionaldehyde group, or a butyl aldehyde group at the other end of the non-peptidyl linker.
양쪽 말단에 히드록시 반응기를 갖는 폴리에틸렌 글리콜을 비펩타이드성 중합체로 이용하는 경우에는 공지의 화학반응에 의해 상기 히드록시기를 상기 다양한 반응기로 활성화하거나, 상업적으로 입수 가능한 변형된 반응기를 갖는 폴리에틸렌 글리콜을 이용하여 본 발명의 지속형 단백질 결합체를 제조할 수 있다.When polyethylene glycol having a hydroxyl reactive group at both ends is used as a non-peptidyl polymer, the hydroxyl group can be activated into the various reactive groups by a known chemical reaction, or a commercially available polyethylene glycol having a modified reactive group is used. The long-acting protein conjugate of the invention can be prepared.
하나의 구체적인 실시 형태에서 상기 비펩타이드성 중합체는 Z의 시스테인 잔기, 보다 구체적으로 시스테인의 -SH 기에 연결되는 것일 수 있으나, 이에 제한되지 않는다. In one specific embodiment, the non-peptidyl polymer may be linked to a cysteine residue of Z, more specifically, a -SH group of cysteine, but is not limited thereto.
예컨대, 상기 Z에 해당하는 펩타이드에서 10번 시스테인 잔기, 13번 시스테인 잔기, 15번 시스테인 잔기, 17번 시스테인 잔기, 19번 시스테인 잔기, 21번 시스테인 잔기, 24번 시스테인 잔기, 28번 시스테인 잔기, 29번 시스테인 잔기, 30번 시스테인 잔기, 31번 시스테인 잔기, 40번 시스테인 잔기, 또는 41번 시스테인 잔기에 상기 비펩타이드성 중합체가 연결된 것일 수 있으나, 특별히 이에 제한되지 않는다. For example, in the peptide corresponding to Z, cysteine residue 10, cysteine 13, cysteine 15, cysteine 17, cysteine 19, cysteine 21, cysteine 24, cysteine 28, 29 The non-peptidyl polymer may be linked to cysteine residue No. 30, cysteine residue 30, cysteine 31, cysteine 40, or cysteine 41, but is not particularly limited thereto.
구체적으로, 상기 시스테인 잔기의 -SH 기에 비펩타이드성 중합체의 반응기가 연결될 수 있으며, 반응기에 대해서는 앞서 기술한 내용이 모두 적용된다. 만약, 말레이미드-PEG-알데히드를 사용하는 경우, 말레이미드 기는 Z의 -SH 기와 티오에테르(thioether) 결합으로 연결하고, 알데히드기는 Fc, 구체적으로 면역글로불린 Fc의 -NH2기와 환원적 아민화 반응을 통해 연결할 수 있으나, 이에 제한되지 않으며, 이는 하나의 일례에 해당한다. Specifically, a reactive group of the non-peptidyl polymer may be linked to the -SH group of the cysteine residue, and all of the above descriptions apply to the reactive group. If maleimide-PEG-aldehyde is used, the maleimide group is linked to the -SH group of Z by a thioether bond, and the aldehyde group is Fc, specifically, the -NH 2 group of immunoglobulin Fc reductive amination reaction may be connected through, but is not limited thereto, and this corresponds to one example.
이와 같은 환원적 알킬화를 통하여 비펩타이드성 중합체인 PEG의 한쪽 말단에 위치한 산소 원자에 면역글로불린 Fc 영역의 N-말단 아미노기가 -CH2CH2CH2-의 구조를 가지는 링커 작용기를 통해 서로 연결되어, -PEG-O-CH2CH2CH2NH-면역글로불린 Fc와 같은 구조를 형성할 수 있고, 티오에테르 결합을 통하여 PEG의 한쪽 말단이 일반식 1 또는 서열번호 1 내지 102 중 어느 하나의 서열을 포함하는 펩타이드의 시스테인에 위치한 황 원자에 연결된 구조를 형성할 수 있다. 상술한 티오에테르 결합은
Figure PCTKR2021009210-appb-img-000002
의 구조를 포함할 수 있다.Through such reductive alkylation, the N-terminal amino group of the immunoglobulin Fc region is linked to an oxygen atom located at one end of PEG, a non-peptidyl polymer, through a linker functional group having the structure -CH 2 CH 2 CH 2 - , -PEG-O-CH 2 CH 2 CH 2 NH-Can form the same structure as immunoglobulin Fc, and one end of PEG through a thioether bond is a sequence of any one of Formula 1 or SEQ ID NOs: 1 to 102 It can form a structure linked to the sulfur atom located at the cysteine of the peptide containing. The above-mentioned thioether bond is
Figure PCTKR2021009210-appb-img-000002
may contain the structure of
그러나, 상술한 예에 특별히 제한되는 것은 아니며, 이는 하나의 일례에 해당한다.However, it is not particularly limited to the above-described example, and this corresponds to one example.
다른 구체적인 실시 형태에서 상기 비펩타이드성 중합체는 Z의 라이신 잔기, 보다 구체적으로, 라이신의 아미노기에 연결되는 것일 수 있으나, 이에 제한되지 않는다.In another specific embodiment, the non-peptidyl polymer may be linked to a lysine residue of Z, more specifically, an amino group of lysine, but is not limited thereto.
또한, 상기 결합체에서, 비펩타이드성 중합체의 반응기가 면역글로불린 Fc 영역의 N-말단에 위치한 -NH2와 연결된 것일 수 있으나, 이는 하나의 일례에 해당한다.In addition, in the conjugate, the reactive group of the non-peptidyl polymer may be linked to -NH 2 located at the N-terminus of the immunoglobulin Fc region, but this corresponds to one example.
또한, 상기 결합체에서, 펩타이드는 반응기를 갖는 링커와 C-말단을 통해 연결될 수 있으나, 이는 하나의 일례에 해당한다.In addition, in the conjugate, the peptide may be linked to a linker having a reactive group through the C-terminus, but this corresponds to one example.
본 발명에서 "C-말단"은, 펩타이드의 카르복시 말단을 의미하는 것으로, 본 발명의 목적상 링커와 결합할 수 있는 위치를 말한다. 그 예로, 이에 제한되지는 않으나, C-말단의 최말단 아미노산 잔기뿐만 아니라 C-말단 주위의 아미노산 잔기를 모두 포함할 수 있으며, 구체적으로는 최말단으로부터 첫 번째 내지 20 번째의 아미노산 잔기를 포함할 수 있으나, 이에 제한되지 않는다.In the present invention, "C-terminus" refers to the carboxy terminus of a peptide, and refers to a position capable of binding to a linker for the purpose of the present invention. For example, but not limited thereto, it may include all amino acid residues around the C-terminus as well as the most terminal amino acid residue at the C-terminus, and specifically includes the first to 20th amino acid residues from the most terminal. can, but is not limited thereto.
한편, 상기 Fc는 면역글로불린 Fc 영역일 수 있으며, 보다 구체적으로 상기 면역글로불린 Fc 영역은 IgG 유래 일 수 있으나, 특별히 이에 제한되지 않는다. Meanwhile, the Fc may be an immunoglobulin Fc region, and more specifically, the immunoglobulin Fc region may be derived from IgG, but is not particularly limited thereto.
본 발명에서, "면역글로불린 Fc 영역"은, 면역글로불린의 중쇄와 경쇄 가변영역을 제외한, 중쇄 불변영역 2(CH2) 및/또는 중쇄 불변영역 3(CH3)부분을 포함하는 부위를 의미한다. 상기 면역글로불린 Fc 영역은 본 발명의 결합체의 모이어티를 이루는 일 구성일 수 있다. 상기 면역글로불린 Fc 영역은 “면역글로불린 Fc 단편”과 혼용되어 사용될 수 있다.As used herein, the term "immunoglobulin Fc region" refers to a region including heavy chain constant region 2 (CH2) and/or heavy chain constant region 3 (CH3), excluding the heavy and light chain variable regions of immunoglobulin. The immunoglobulin Fc region may be one component constituting a moiety of the conjugate of the present invention. The immunoglobulin Fc region may be used interchangeably with “immunoglobulin Fc fragment”.
본 명세서에서 Fc 영역이라고 하면 면역글로불린의 파파인 소화에서 얻는 천연형 서열뿐 아니라 그 유도체, 예컨대 천연 서열 중의 하나 이상의 아미노산 잔기가 결실, 삽입, 비보전적 또는 보전적 치환 또는 이들의 조합에 의하여 변환되어 천연형과 상이하게 된 서열까지 망라하여 포함된다.　In the present specification, referring to the Fc region, not only the native sequence obtained from papain digestion of immunoglobulin, but also its derivatives, such as one or more amino acid residues in the native sequence, are converted by deletion, insertion, non-conservative or conservative substitution, or a combination thereof, resulting in a native natural sequence. It encompasses and includes sequences that differ from the type.
상기 Fc 는, 2개의 폴리펩타이드 사슬이 이황화결합으로 연결되어 있는 구조이며, 상기 두 사슬 중 한 사슬의 질소 원자를 통해서만 연결되어 있는 구조일 수 있으나, 이에 제한되지 않는다. 상기 질소 원자를 통한 연결은 리신의 입실론 아미노 원자나 N-말단 아미노기에 환원적 아민화를 통하여 연결될 수 있다.The Fc has a structure in which two polypeptide chains are linked by a disulfide bond, and may have a structure in which only one of the two chains is linked through a nitrogen atom, but is not limited thereto. The linkage via the nitrogen atom may be linked via reductive amination to the epsilon amino atom or the N-terminal amino group of lysine.
환원적 아민화 반응이란 반응물의 아민기 또는 아미노기가 다른 반응물의 알데히드 (즉, 환원적 아민화가 가능한 작용기)와 반응하여 아민을 생성한 다음, 환원 반응에 의해 아민 결합을 형성시키는 반응을 의미하여, 당해 기술 분야에 널리 알려져 있는 유기합성 반응이다.Reductive amination reaction refers to a reaction in which an amine group or an amino group of a reactant reacts with an aldehyde (that is, a functional group capable of reductive amination) of another reactant to form an amine, and then forms an amine bond by a reduction reaction, It is an organic synthesis reaction well known in the art.
하나의 구체예로, 상기 Fc는 그 N-말단 프롤린의 질소 원자를 통하여 연결된 것일 수 있으나, 이에 제한되지 않는다.In one embodiment, the Fc may be linked through the nitrogen atom of its N-terminal proline, but is not limited thereto.
상기 면역글로불린 Fc 영역은 본 발명의 화학식 1의 결합체의 모이어티를 이루는 일 구성으로, 구체적으로, 상기 화학식 1에서 Fc에 해당할 수 있다.The immunoglobulin Fc region is one component constituting a moiety of the conjugate of Formula 1 of the present invention, and specifically, may correspond to Fc in Formula 1 above.
이러한 면역글로불린 Fc 영역은 중쇄 불변영역에 힌지(hinge) 부분을 포함할 수 있으나, 이에 제한되는 것은 아니다. The immunoglobulin Fc region may include a hinge region in the heavy chain constant region, but is not limited thereto.
본 발명에서, 면역글로불린 Fc 영역은 N-말단에 특정 힌지 서열을 포함할 수 있다. In the present invention, the immunoglobulin Fc region may include a specific hinge sequence at the N-terminus.
본 발명의 용어, "힌지 서열"은 중쇄에 위치하여 이황화결합(inter disulfide bond)를 통하여 면역글로불린 Fc 영역의 이량체를 형성하는 부위를 의미한다. As used herein, the term “hinge sequence” refers to a region that is located on a heavy chain and forms a dimer of an immunoglobulin Fc region through an inter disulfide bond.
본 발명에서, 상기 힌지 서열은 하기의 아미노산 서열을 갖는 힌지 서열 중 일부가 결실되어 하나의 시스테인 잔기만을 갖도록 변이된 것일 수 있으나, 이에 제한되지 않는다:In the present invention, the hinge sequence may be mutated to have only one cysteine residue by deleting a portion of the hinge sequence having the following amino acid sequence, but is not limited thereto:
Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser-Cys-Pro(서열번호 118).Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser-Cys-Pro (SEQ ID NO: 118).
상기 힌지 서열은 서열번호 118의 힌지 서열 중 8번째 또는 11번째 시스테인 잔기가 결실되어 하나의 시스테인 잔기만을 포함하는 것일 수 있다. 본 발명의 힌지 서열은 하나의 시스테인 잔기만을 포함하는, 3 내지 12개의 아미노산으로 구성된 것일 수 있으나, 이에 제한되지 않는다. 보다 구체적으로, 본 발명의 힌지 서열은 다음과 같은 서열을 가질 수 있다: Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Pro-Ser-Cys-Pro(서열번호 119), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser-Pro(서열번호 120), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser(서열번호 121), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Pro(서열번호 122), Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser(서열번호 123), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys(서열번호 124), Glu-Lys-Tyr-Gly-Pro-Pro-Cys(서열번호 125), Glu-Ser-Pro-Ser-Cys-Pro(서열번호 126), Glu-Pro-Ser-Cys-Pro(서열번호 127), Pro-Ser-Cys-Pro(서열번호 128), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Ser-Cys-Pro(서열번호 129), Lys-Tyr-Gly-Pro-Pro-Pro-Ser-Cys-Pro(서열번호 130), Glu-Ser-Lys-Tyr-Gly-Pro-Ser-Cys-Pro(서열번호 131), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys(서열번호 132), Lys-Tyr-Gly-Pro-Pro-Cys-Pro(서열번호 133), Glu-Ser-Lys-Pro-Ser-Cys-Pro(서열번호 134), Glu-Ser-Pro-Ser-Cys-Pro(서열번호 135), Glu-Pro-Ser-Cys(서열번호 136), Ser-Cys-Pro(서열번호 137). The hinge sequence may include only one cysteine residue by deleting the 8th or 11th cysteine residue in the hinge sequence of SEQ ID NO: 118. The hinge sequence of the present invention may be composed of 3 to 12 amino acids, including only one cysteine residue, but is not limited thereto. More specifically, the hinge sequence of the present invention may have the following sequences: Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Pro-Ser-Cys-Pro (SEQ ID NO: 119), Glu-Ser- Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser-Pro (SEQ ID NO: 120), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser (SEQ ID NO: 121), Glu- Ser-Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Pro (SEQ ID NO: 122), Lys-Tyr-Gly-Pro-Pro-Cys-Pro-Ser (SEQ ID NO: 123), Glu-Ser-Lys- Tyr-Gly-Pro-Pro-Cys (SEQ ID NO: 124), Glu-Lys-Tyr-Gly-Pro-Pro-Cys (SEQ ID NO: 125), Glu-Ser-Pro-Ser-Cys-Pro (SEQ ID NO: 126) , Glu-Pro-Ser-Cys-Pro (SEQ ID NO: 127), Pro-Ser-Cys-Pro (SEQ ID NO: 128), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Ser-Cys-Pro (SEQ ID NO: 128) No. 129), Lys-Tyr-Gly-Pro-Pro-Pro-Ser-Cys-Pro (SEQ ID NO: 130), Glu-Ser-Lys-Tyr-Gly-Pro-Ser-Cys-Pro (SEQ ID NO: 131), Glu-Ser-Lys-Tyr-Gly-Pro-Pro-Cys (SEQ ID NO: 132), Lys-Tyr-Gly-Pro-Pro-Cys-Pro (SEQ ID NO: 133), Glu-Ser-Lys-Pro-Ser- Cys-Pro (SEQ ID NO: 134), Glu-Ser-Pro-Ser-Cys-Pro (SEQ ID NO: 135), Glu-Pro-Ser-Cys (SEQ ID NO: 136), Ser-Cys-Pro (SEQ ID NO: 137).
더욱 구체적으로는 상기 힌지 서열은 서열번호 128(Pro-Ser-Cys-Pro)또는 서열번호 137(Ser-Cys-Pro)의 아미노산 서열을 포함하는 것일 수 있으나, 이에 제한되지 않는다. More specifically, the hinge sequence may include the amino acid sequence of SEQ ID NO: 128 (Pro-Ser-Cys-Pro) or SEQ ID NO: 137 (Ser-Cys-Pro), but is not limited thereto.
본 발명의 면역글로불린 Fc 영역은 힌지 서열의 존재로 면역글로불린 Fc 사슬 두 분자가 이량체를 형성한 형태일 수 있고, 또한, 본 발명의 화학식 1의 결합체는 링커의 일 말단이 이량체의 면역글로불린 Fc 영역의 한 사슬에 연결된 형태일 수 있으나, 이에 제한되는 것은 아니다.The immunoglobulin Fc region of the present invention may be in a form in which two immunoglobulin Fc chain molecules form a dimer due to the presence of a hinge sequence. In addition, in the conjugate of Formula 1, one end of the linker is a dimer immunoglobulin It may be in a form linked to one chain of the Fc region, but is not limited thereto.
본 발명의 용어, "N-말단"은 단백질 또는 폴리펩티드의 아미노 말단을 의미하는 것으로, 아미노 말단의 최말단, 또는 최말단으로부터 1개, 2개, 3개, 4개, 5개, 6개, 7개, 8개, 9개, 또는 10개 이상의 아미노산까지 포함하는 것일 수 있다. 본 발명의 면역글로불린 Fc 영역은 힌지 서열을 N-말단에 포함할 수 있으나, 이에 제한되지 않는다.As used herein, the term “N-terminus” refers to the amino terminus of a protein or polypeptide, and 1, 2, 3, 4, 5, 6, It may include up to 7, 8, 9, or 10 or more amino acids. The immunoglobulin Fc region of the present invention may include a hinge sequence at the N-terminus, but is not limited thereto.
또한 본 발명의 면역글로불린 Fc 영역은 천연형과 실질적으로 동등하거나 향상된 효과를 갖는 한, 면역 글로불린의 중쇄와 경쇄 가변영역만을 제외하고, 일부 또는 전체 중쇄 불변영역 1(CH1) 및/또는 경쇄불변영역 1(CL1)을 포함하는 확장된 Fc 영역일 수 있다. 또한, CH2 및/또는 CH3에 해당하는 상당히 긴 일부 아미노산 서열이 제거된 영역일 수도 있다. In addition, as long as the immunoglobulin Fc region of the present invention has substantially the same or improved effect as the native type, part or all of the heavy chain constant region 1 (CH1) and/or the light chain constant region except for only the heavy and light chain variable regions of immunoglobulin 1 (CL1) may be an extended Fc region. Also, it may be a region in which some fairly long amino acid sequences corresponding to CH2 and/or CH3 have been removed.
예컨대, 본 발명의 면역글로불린 Fc 영역은 1) CH1 도메인, CH2 도메인, CH3 도메인 및 CH4 도메인, 2) CH1 도메인 및 CH2 도메인, 3) CH1 도메인 및 CH3 도메인, 4) CH2 도메인 및 CH3 도메인, 5) CH1 도메인, CH2 도메인, CH3 도메인 및 CH4 도메인 중 1개 또는 2개의 이상의 도메인과 면역글로불린 힌지 영역(또는 힌지 영역의 일부)와의 조합, 6) 중쇄 불변 영역 각 도메인과 경쇄 불변영역의 이량체일 수 있다. 그러나, 이에 제한되는 것은 아니다. For example, the immunoglobulin Fc region of the present invention comprises 1) a CH1 domain, a CH2 domain, a CH3 domain and a CH4 domain, 2) a CH1 domain and a CH2 domain, 3) a CH1 domain and a CH3 domain, 4) a CH2 domain and a CH3 domain, 5) a combination of one or two or more domains of CH1 domain, CH2 domain, CH3 domain and CH4 domain with an immunoglobulin hinge region (or a part of the hinge region), 6) heavy chain constant region Each domain may be a dimer of a light chain constant region . However, the present invention is not limited thereto.
또한, 하나의 구체예로서, 상기 면역글로불린 Fc 영역은 이량체 형태 (dimeric form)일 수 있으며, 이량체 형태의 하나의 Fc 영역에 Z 한 분자가 공유결합적으로 연결될 수 있으며, 이때 상기 면역글로불린 Fc와 Z는 에틸렌글리콜 반복 단위를 함유하는 링커를 통하여 공유결합으로 서로 연결될 수 있다. 한편, 이량체 형태의 하나의 Fc 영역에 Z 두 분자가 대칭적으로 결합하는 것 역시 가능하다. 이때 상기 면역글로불린 Fc와 Z는 에틸렌글리콜 반복 단위를 함유하는 링커에 의해 서로 연결될 수 있다. 그러나, 상기 기술된 예에 제한되는 것은 아니다. 하나의 구체예에서, Z는 이러한 Fc 영역 이량체 Fc의 두 폴리펩타이드 사슬 중 하나의 폴리펩타이드 사슬에만 링커 Lx를 통하여 공유결합으로 연결되어 있을 수 있다. 그 예로, 이러한 Fc 영역 이량체 Fc의 두 폴리펩타이드 사슬 중 Z가 연결된 하나의 폴리펩타이드 사슬에는 한 분자의 Z만이 Lx를 통하여 공유결합적으로 연결되어 있을 수 있다. 그 예로 상기 Fc는 동종이량체(homodimer)일 수 있다.In addition, in one embodiment, the immunoglobulin Fc region may be in a dimeric form, and one molecule of Z may be covalently linked to one Fc region in a dimeric form, in which case the immunoglobulin Fc and Z may be covalently linked to each other through a linker containing an ethylene glycol repeating unit. On the other hand, it is also possible for two molecules of Z to bind symmetrically to one Fc region in a dimer form. In this case, the immunoglobulin Fc and Z may be linked to each other by a linker containing an ethylene glycol repeating unit. However, it is not limited to the examples described above. In one embodiment, Z may be covalently linked via a linker Lx to only one of the two polypeptide chains of the Fc region dimer Fc. For example, among the two polypeptide chains of the Fc region dimer Fc, only one molecule of Z may be covalently linked via Lx to one polypeptide chain to which Z is linked. For example, the Fc may be a homodimer.
다른 구체예에서, 상기 면역글로불린 Fc 영역인 Fc는 두 개의 폴리펩타이드 사슬로 이루어진 이량체이며, Lx의 한 말단이 상기 두 폴리펩타이드 사슬 중 하나의 폴리펩티드 사슬에만 연결되어 있는 것일 수 있으나, 이에 제한되지 않는다. In another embodiment, the immunoglobulin Fc region Fc is a dimer consisting of two polypeptide chains, and one end of Lx may be connected to only one of the two polypeptide chains, but is not limited thereto. does not
또한, 본 발명의 면역글로불린 Fc 영역은 천연형 아미노산 서열뿐만 아니라 이의 서열 유도체를 포함한다. 아미노산 서열 유도체란 천연 아미노산 서열 중의 하나 이상의 아미노산 잔기가 결실, 삽입, 비보전적 또는 보전적 치환 또는 이들의 조합에 의하여 상이한 서열을 가지는 것을 의미한다.In addition, the immunoglobulin Fc region of the present invention includes a native amino acid sequence as well as a sequence derivative thereof. An amino acid sequence derivative means that one or more amino acid residues in a natural amino acid sequence have a different sequence by deletion, insertion, non-conservative or conservative substitution, or a combination thereof.
예를 들면, IgG Fc의 경우 결합에 중요하다고 알려진 214 내지 238, 297 내지 299, 318 내지 322 또는 327 내지 331번 아미노산 잔기들이 변형을 위해 적당한 부위로서 이용될 수 있다.For example, in the case of IgG Fc, amino acid residues 214 to 238, 297 to 299, 318 to 322, or 327 to 331 known to be important for binding may be used as suitable sites for modification.
또한, 이황화 결합을 형성할 수 있는 부위가 제거되거나, 천연형 Fc에서 N-말단의 몇몇 아미노산이 제거되거나 또는 천연형 Fc의 N-말단에 메티오닌 잔기가 부가될 수도 있는 등 다양한 종류의 유도체가 가능하다. 또한, 이펙터 기능을 없애기 위해 보체결합부위, 예로 C1q 결합부위가 제거될 수도 있고, ADCC (antibody dependent cell mediated cytotoxicity) 부위가 제거될 수도 있다. 이러한 면역글로불린 Fc 영역의 서열 유도체를 제조하는 기술은 국제특허공개 제WO 97/34631호, 국제특허공개 제96/32478호 등에 개시되어 있다.In addition, various types of derivatives are possible, such as a site capable of forming a disulfide bond is removed, some amino acids at the N-terminus of native Fc are removed, or a methionine residue may be added to the N-terminus of native Fc do. In addition, in order to eliminate the effector function, the complement binding site, eg, the C1q binding site, may be removed, or the ADCC (antibody dependent cell mediated cytotoxicity) site may be removed. Techniques for preparing such an immunoglobulin Fc region sequence derivative are disclosed in International Patent Publication Nos. WO 97/34631 and International Patent Publication No. 96/32478.
분자의 활성을 전체적으로 변경시키지 않는 단백질 및 펩타이드에서의 아미노산 교환은 당해 분야에 공지되어 있다 (H.Neurath, R.L.Hill, The Proteins, Academic Press, New York, 1979). 가장 통상적으로 일어나는 교환은 아미노산 잔기 Ala/Ser, Val/Ile, Asp/Glu, Thr/Ser, Ala/Gly, Ala/Thr, Ser/Asn, Ala/Val, Ser/Gly, Thy/Phe, Ala/Pro, Lys/Arg, Asp/Asn, Leu/Ile, Leu/Val, Ala/Glu, Asp/Gly 간의 교환이다. 경우에 따라서는 인산화(phosphorylation), 황화(sulfation), 아크릴화(acrylation), 당화(glycosylation), 메틸화(methylation), 파네실화(farnesylation), 아세틸화(acetylation) 및 아미드화(amidation) 등으로 수식(modification)될 수도 있다.Amino acid exchanges in proteins and peptides that do not entirely alter the activity of the molecule are known in the art (H.Neurath, R.L.Hill, The Proteins, Academic Press, New York, 1979). The most common exchanges are amino acid residues Ala/Ser, Val/Ile, Asp/Glu, Thr/Ser, Ala/Gly, Ala/Thr, Ser/Asn, Ala/Val, Ser/Gly, Thy/Phe, Ala/ It is an exchange between Pro, Lys/Arg, Asp/Asn, Leu/Ile, Leu/Val, Ala/Glu, Asp/Gly. In some cases, the formula (phosphorylation, sulfation, acrylation, glycosylation, methylation, farnesylation, acetylation, amidation, etc.) may be modified).
상기 기술한 Fc 유도체는 본 발명의 Fc 영역과 동등한 생물학적 활성을 나타내며 Fc 영역의 열, pH 등에 대한 구조적 안정성을 증대시킨 것일 수 있다. The above-described Fc derivative may exhibit biological activity equivalent to that of the Fc region of the present invention, and may have increased structural stability to heat, pH, etc. of the Fc region.
또한, 이러한 Fc 영역은 인간, 소, 염소, 돼지, 마우스, 래빗, 햄스터, 랫트 또는 기니아 픽 등의 동물의 생체 내에서 분리한 천연형으로부터 얻어질 수도 있고, 형질전환된 동물세포 또는 미생물로부터 얻어진 재조합형 또는 이의 유도체일 수 있다. 여기서, 천연형으로부터 획득하는 방법은 전체 면역글로불린을 인간 또는 동물의 생체로부터 분리한 후, 단백질 분해효소를 처리하여 획득하는 방법일 수 있다. 파파인을 처리할 경우에는 Fab 및 Fc로 절단되고, 펩신을 처리할 경우에는 pF'c 및 F(ab)2로 절단된다. 이를 크기 배제 크로마토그래피 (size-exclusion chromatography) 등을 이용하여 Fc 또는 pF'c를 분리할 수 있다. 더 구체적인 실시 형태에서는 인간 유래의 Fc 영역을 미생물로부터 수득한 재조합형 면역글로불린 Fc 영역이다.In addition, the Fc region may be obtained from a native type isolated in vivo from animals such as humans, cows, goats, pigs, mice, rabbits, hamsters, rats or guinea pigs, or obtained from transformed animal cells or microorganisms. It may be recombinant or a derivative thereof. Here, the method of obtaining from the native type may be a method of obtaining whole immunoglobulin by isolating it from a living body of a human or animal and then treating it with a proteolytic enzyme. When treated with papain, it is cleaved into Fab and Fc, and when treated with pepsin, it is cleaved into pF'c and F(ab) 2 . In this case, Fc or pF'c may be separated using size-exclusion chromatography or the like. In a more specific embodiment, it is a recombinant immunoglobulin Fc region obtained by obtaining a human-derived Fc region from a microorganism.
또한, 면역글로불린 Fc 영역은 천연형 당쇄, 천연형에 비해 증가된 당쇄, 천연형에 비해 감소한 당쇄 또는 당쇄가 제거된 형태일 수 있다. 이러한 면역글로불린 Fc 당쇄의 증감 또는 제거에는 화학적 방법, 효소학적 방법 및 미생물을 이용한 유전 공학적 방법과 같은 통상적인 방법이 이용될 수 있다. 여기서, Fc에서 당쇄가 제거된 면역글로불린 Fc 영역은 보체(c1q)와의 결합력이 현저히 저하되고, 항체-의존성 세포독성 또는 보체-의존성 세포 독성이 감소 또는 제거되므로, 생체 내에서 불필요한 면역 반응을 유발하지 않는다. 이런 점에서 약물의 캐리어로서의 본래의 목적에 보다 부합하는 형태는 당쇄가 제거되거나 비당쇄화된 면역글로불린 Fc 영역이라 할 것이다.In addition, the immunoglobulin Fc region may have a native sugar chain, an increased sugar chain compared to the native type, a decreased sugar chain compared to the native type, or a form in which the sugar chain is removed. Conventional methods such as chemical methods, enzymatic methods, and genetic engineering methods using microorganisms may be used for the increase or decrease or removal of such immunoglobulin Fc sugar chains. Here, the immunoglobulin Fc region from which the sugar chains are removed from the Fc has significantly reduced binding to complement (c1q) and reduced or eliminated antibody-dependent cytotoxicity or complement-dependent cytotoxicity, so that unnecessary immune responses in vivo are not induced. does not In this respect, a form more suitable for the original purpose as a drug carrier will be an immunoglobulin Fc region in which sugar chains are removed or non-glycosylated.
본 발명에서 "당쇄의 제거(Deglycosylation)"는 효소로 당을 제거한 Fc 영역을 말하며, 비당쇄화(Aglycosylation)는 원핵동물, 더 구체적인 실시 형태에서는 대장균에서 생산하여 당쇄화되지 않은 Fc 영역을 의미한다.In the present invention, "deglycosylation" refers to an Fc region from which sugars have been removed with an enzyme, and aglycosylation refers to an Fc region that is not glycosylated by production in prokaryotes, in a more specific embodiment, in E. coli. .
한편, 면역글로불린 Fc 영역은 인간 또는 소, 염소, 돼지, 마우스, 래빗, 햄스터, 랫트, 기니아 픽 등의 동물기원일 수 있으며, 더 구체적인 실시 형태에서는 인간기원이다.Meanwhile, the immunoglobulin Fc region may be of human or animal origin, such as cattle, goats, pigs, mice, rabbits, hamsters, rats, and guinea pigs, and in a more specific embodiment, it is of human origin.
또한, 면역글로불린 Fc 영역은 IgG, IgA, IgD, IgE, IgM 유래 또는 이들의 조합(combination) 또는 이들의 혼성(hybrid)에 의한 Fc 영역일 수 있다. 더 구체적인 실시 형태에서는 인간 혈액에 가장 풍부한 IgG 또는 IgM유래이며, 보다 더 구체적인 실시 형태에서는 리간드 결합 단백질의 반감기를 향상시키는 것으로 공지된 IgG 유래이다. 더욱 더 구체적인 실시 형태에서 상기 면역글로불린 Fc 영역은 IgG4 Fc 영역이며, 가장 구체적인 실시 형태에서 상기 면역글로불린 Fc 영역은 인간 IgG4 유래의 비-당쇄화된 Fc 영역이나, 이에 제한되는 것은 아니다.In addition, the immunoglobulin Fc region may be an Fc region derived from IgG, IgA, IgD, IgE, or IgM, or a combination or hybrid thereof. In a more specific embodiment, it is derived from IgG or IgM, which is most abundant in human blood, and in a more specific embodiment, it is derived from IgG, which is known to enhance the half-life of ligand binding proteins. In an even more specific embodiment, the immunoglobulin Fc region is an IgG4 Fc region, and in the most specific embodiment, the immunoglobulin Fc region is a non-glycosylated Fc region derived from human IgG4, but is not limited thereto.
또한, 하나의 구체적인 실시 형태에서, 면역글로불린 Fc 절편은 인간 IgG4 Fc의 단편으로서, 각 단량체(monomer)의 3번 아미노산인 시스테인 사이의 이황화 결합(inter-chain 형태)을 통해 2개의 단량체가 연결된 동종이량체(homodimer) 형태일 수 있으며, 이 때 동종이량체의 각 단량체는 독립적으로 35번 및 95번의 시스테인 간의 내부의 이황화 결합 및 141번 및 199번의 시스테인 간의 내부의 이황화 결합, 즉 2개의 내부의 이황화 결합(intra-chain 형태)을 가지거나/가질 수 있다. 각 단량체의 아미노산 수는 221개의 아미노산으로 구성될 수 있므며, 동종이량체를 형성하는 아미노산은 전체 442개의 아미노산으로 이루어질 수 있으나, 이에 제한되지 않는다. 구체적으로 면역글로불린 Fc 절편은 서열번호 138의 아미노산 서열 (221개의 아미노산으로 구성됨)을 갖는 단량체 2개가 각 단량체의 3번 아미노산인 시스테인 사이에 이황화 결합을 통해 동종이량체를 형성하고, 상기 동종이량체의 단량체는 각각 독립적으로 35번 및 95번의 시스테인 간의 내부의 이황화 결합 및 141번 및 199번의 시스테인 간의 내부의 이황화 결합을 형성하는 것일 수 있으나, 이에 제한되지 않는다.Further, in one specific embodiment, the immunoglobulin Fc fragment is a fragment of human IgG4 Fc, and is a homologous type in which two monomers are linked through a disulfide bond (inter-chain form) between cysteine, amino acid 3 of each monomer. It may be in the form of a dimer, wherein each monomer of the homodimer is independently an internal disulfide bond between cysteines at positions 35 and 95 and an internal disulfide bond between cysteines at positions 141 and 199, that is, two internal It may have/have disulfide bonds (intra-chain form). The number of amino acids of each monomer may consist of 221 amino acids, and the amino acids forming the homodimer may consist of a total of 442 amino acids, but is not limited thereto. Specifically, in the immunoglobulin Fc fragment, two monomers having the amino acid sequence of SEQ ID NO: 138 (consisting of 221 amino acids) form a homodimer through a disulfide bond between cysteine, the 3rd amino acid of each monomer, and the homodimer The monomers of may each independently form an internal disulfide bond between cysteines at positions 35 and 95 and an internal disulfide bond between cysteines at positions 141 and 199, but is not limited thereto.
상기 화학식 1의 Fc는 서열번호 138의 아미노산 서열인 단량체를 포함하는 것일 수 있으며, 상기 Fc는 서열번호 138의 아미노산 서열의 단량체의 동종이량체일 수 있으나, 이에 제한되지 않는다.The Fc of Formula 1 may include a monomer having the amino acid sequence of SEQ ID NO: 138, and the Fc may be a homodimer of the monomer having the amino acid sequence of SEQ ID NO: 138, but is not limited thereto.
하나의 예로, 면역글로불린 Fc 영역은 서열번호 139의 아미노산 서열 (442개의 아미노산으로 구성됨)을 포함하는 동종이량체일 수 있으나, 이에 제한되지 않는다.As an example, the immunoglobulin Fc region may be a homodimer comprising the amino acid sequence of SEQ ID NO: 139 (consisting of 442 amino acids), but is not limited thereto.
한편, 본 발명에서 면역글로불린 Fc 영역과 관련된 "조합(combination)"이란 이량체 또는 다량체를 형성할 때, 동일 기원 단쇄 면역글로불린 Fc 영역을 암호화하는 폴리펩타이드가 상이한 기원의 단쇄 폴리펩타이드와 결합을 형성하는 것을 의미한다. 즉, IgG Fc, IgA Fc, IgM Fc, IgD Fc 및 IgE의 Fc 단편으로 이루어진 그룹으로부터 선택된 2개 이상의 단편으로부터 이량체 또는 다량체의 제조가 가능하다.Meanwhile, in the present invention, the term "combination" with respect to an immunoglobulin Fc region means that when a dimer or multimer is formed, a polypeptide encoding a single-chain immunoglobulin Fc region of the same origin binds to a single-chain polypeptide of a different origin. means to form That is, it is possible to prepare a dimer or multimer from two or more fragments selected from the group consisting of IgG Fc, IgA Fc, IgM Fc, IgD Fc and IgE Fc fragment.
본 발명에서 "하이브리드(hybrid)"란 단쇄의 면역글로불린 불변영역 내에 2개 이상의 상이한 기원의 면역글로불린 Fc 단편에 해당하는 서열이 존재함을 의미하는 용어이다. 본 발명의 경우 여러 형태의 하이브리드가 가능하다. 즉, IgG Fc, IgM Fc, IgA Fc, IgE Fc 및 IgD Fc의 CH1, CH2, CH3 및 CH4로 이루어진 그룹으로부터 1개 내지 4개 도메인으로 이루어진 도메인의 하이브리드가 가능하며, 힌지를 포함할 수 있다.In the present invention, "hybrid" is a term meaning that sequences corresponding to immunoglobulin Fc fragments of two or more different origins exist in a single-chain immunoglobulin constant region. In the case of the present invention, various types of hybrids are possible. That is, a hybrid of domains consisting of 1 to 4 domains from the group consisting of CH1, CH2, CH3 and CH4 of IgG Fc, IgM Fc, IgA Fc, IgE Fc and IgD Fc is possible, and may include a hinge.
한편, IgG 역시 IgG1, IgG2, IgG3 및 IgG4의 서브클래스로 나눌 수 있고 본 발명에서는 이들의 조합 또는 이들의 혼성화도 가능하다. 구체적으로는 IgG2 및 IgG4 서브클래스이며, 가장 구체적으로는 보체 의존적 독성(CDC, Complement dependent cytotoxicity)과 같은 이펙터 기능(effector function)이 거의 없는 IgG4의 Fc 절편이다.On the other hand, IgG can also be divided into subclasses of IgG1, IgG2, IgG3 and IgG4, and in the present invention, a combination thereof or hybridization thereof is also possible. Specifically, they are subclasses of IgG2 and IgG4, and most specifically, an Fc fragment of IgG4 having little effector function such as complement dependent cytotoxicity (CDC).
또한, 상술한 결합체는 효력의 지속성이 천연형 GLP-1, GIP, 혹은 글루카곤에 비해, 또는 Fc가 수식되지 않은 Z에 비해 증가된 것일 수 있으며, 이러한 결합체는 상술한 형태뿐만 아니라, 생분해성 나노파티클에 봉입된 형태 등을 모두 포함하나 이에 제한되지 않는다.In addition, the above-described conjugate may have an increased duration of effect compared to native GLP-1, GIP, or glucagon, or compared to Z in which Fc is not modified. It includes all types of particles and the like, but is not limited thereto.
상기 FXR 작용제는 담즙산 수용체(Bile acid Receptor, BAR)로도 공지되어 있는, 담즙산에 의해 활성화되는 핵 수용체인 FXR의 작용제를 의미하는 것으로, 본 발명의 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체와 병용 요법에 사용되어, 단독에 비해 간 질환을 예방 또는 치료 효과를 개선시킬 수 있는 물질은 제한없이 포함될 수 있다.The FXR agonist refers to an agonist of FXR, a nuclear receptor activated by bile acids, also known as Bile acid Receptor (BAR), for the glucagon receptor, GLP-1 receptor, and GIP receptor of the present invention. Substances having an activity (triple agonist) or long-acting combination thereof and used in combination therapy to improve the effect of preventing or treating liver disease compared to alone may be included without limitation.
FXR은 간, 장 및 신장과 같은 담즙산 대사의 주요 부위에서 발현되며, 이는 조직-특이적 방식으로 담즙산을 포함한 여러 대사 경로에 영향을 미친다. FXR은 간 및 장의 여러 메커니즘을 통해 담즙산 생산억제, 및 제거촉진 작용을 한다고 알려져있다. 또한, FXR 작용제는 간 트리글리세라이드 합성을 감소시켜 지방증을 감소시키고, 간 성상 세포 활성화를 억제하여, 간 섬유증을 감소시키고, FGF15/FGF19 발현(담즙산 대사의 주요 조절제)을 자극하여, 간 인슐린 민감성을 개선시키는 것으로 알려져 있으나, 본 발명의 삼중 활성체 또는 이의 결합체 (3중 작용성 지속형 결합체)와 병용하는 조합물로 이용될 경우 단독에 비해 간 질환의 예방 또는 치료 효과가 개선될 수 있다는 것은 알려진 바 없다.FXR is expressed at key sites of bile acid metabolism, such as the liver, intestine, and kidney, and it affects several metabolic pathways, including bile acids, in a tissue-specific manner. FXR is known to inhibit bile acid production and promote elimination through various mechanisms in the liver and intestine. In addition, FXR agonists reduce hepatic triglyceride synthesis to reduce steatosis, inhibit hepatic stellate cell activation to reduce hepatic fibrosis, stimulate FGF15/FGF19 expression (a major regulator of bile acid metabolism), and thereby reduce hepatic insulin sensitivity. Although it is known to improve no bar
상기 FXR 작용제는 본 발명의 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체)와 함께 병용 요법에 사용되어, 단독에 비해 간 질환을 예방 또는 치료 효과를 개선시킬 수 있는 물질은 제한없이 포함되나, 그 예로 카페스톨 (Cafestol), 케노데옥시콜산 (Chenodeoxycholic acid), 오베티콜산 (Obeticholic acid), 펙사라민 (Fexaramine), GW 4064, PX104, 6E-CDCA (6-ethyl-chedeoxycholic acid), AKN-083 또는 트로피펙서 (Tropifexor), 킬로펙서 (Cilofexor), EDP-305, AGN-242266, AGN-242256, EP-024297, RDX-023, BWL-200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, PX20606, EYP-001, TERN-101, TC-100 또는 INT-2228등 일 수 있다.The FXR agonist is used in combination therapy together with a substance having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) of the present invention. , Substances capable of improving the effect of preventing or treating liver disease compared to alone are included without limitation, for example, cafestol, chenodeoxycholic acid, obeticholic acid, fexara Fexaramine, GW 4064, PX104, 6E-CDCA (6-ethyl-chedeoxycholic acid), AKN-083 or Tropifexor, Cilofexor, EDP-305, AGN-242266, AGN-242256, EP-024297, RDX-023, BWL-200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, It may be PX20606, EYP-001, TERN-101, TC-100, or INT-2228.
상기 ACC 억제제는 지방산 생성 및 대사의 조절에 중요한 효소인 아세틸-CoA 카복실라제 (acetyla-CoA carboxylase)를 억제하는 물질을 의미하며, 본 발명의 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체) 및 상기 FXR 작용제와의 병용 요법에 사용되어, 단독에 비해 간 질환을 예방 또는 치료 효과를 개선시킬 수 있는 물질은 제한없이 포함될 수 있다.The ACC inhibitor refers to a substance that inhibits acetyla-CoA carboxylase, an enzyme important for the regulation of fatty acid production and metabolism, and is active on the glucagon receptor, GLP-1 receptor, and GIP receptor of the present invention A substance having (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) and a substance that can be used in combination therapy with the FXR agonist to prevent or improve the therapeutic effect of liver disease compared to alone may be included without limitation.
상기 ACC 억제제는 본 발명의 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), 및 FXR 작용제와 함께 병용 요법에 사용되어, 단독에 비해 간 질환을 예방 또는 치료 효과를 개선시킬 수 있는 물질은 제한없이 포함되나, 그 예로 CP-640186, (4-피페리디닐)-피페라진 유도체 ((4-piperidinyl)-piperazine derivatives), 1,4-치환 사이클로헥산 유도체 (1,4-disubstituted cyclohexane derivatives), 스피로크로마논 유도체 (Spirochromanone derivatives), 스피로락탐 유도체 (Spirolactam derivatives), 스피로디아민 유도체 (Spirodiamine derivatives), 스피로펜타실아미드 유도체 (Spiropentacylamide derivatives), 슈도펩타이드 피롤리딘다이온 유도체 (Pseudopeptide pyrrolidinedione derivatives), 매크로사이클릭 폴리케톤 유도체 (Macrocyclic polyketone derivatives), 피르소코스타트 (firsocostat)을 포함하는 티오펜 피리미돈 유도체 (Thiophene pyrimidone derivatives), 아미노-옥사졸 유도체 (Amino-oxazole derivatives), 아조벤지미다졸 유도체 (Azobenzimidazole derivatives) 또는 PF-05221304 일 수 있다. 상기 ACC 억제제는 공지의 문헌인 Future Med Chem. 2020 Mar;12(6):533-561 (doi: 10.4155/fmc-2019-0312) 등을 참고할 수 있으며, 상기 문헌은 전체가 본원의 참조로 이용된다.The ACC inhibitor is a substance having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor of the present invention (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), and in combination with an FXR agonist Substances that can be used for therapy and improve the effect of preventing or treating liver disease compared to alone are included without limitation, for example, CP-640186, (4-piperidinyl)-piperazine derivative ((4-piperidinyl) -piperazine derivatives), 1,4-disubstituted cyclohexane derivatives, spirochromanone derivatives, spirolactam derivatives, spirodiamine derivatives, spiropenta Thiophene pyrimidone derivatives including Spiropentacylamide derivatives, Pseudopeptide pyrrolidinedione derivatives, Macrocyclic polyketone derivatives, firsocostat derivatives), amino-oxazole derivatives, azobenzimidazole derivatives or PF-05221304. The ACC inhibitor is a well-known document, Future Med Chem. 2020 Mar; 12(6):533-561 (doi: 10.4155/fmc-2019-0312), etc., which are incorporated herein by reference in their entirety.
본 발명의 조합물, 약학적 조성물 또는 키트는 간 질환의 예방 또는 치료에 사용될 수 있다.The combination, pharmaceutical composition or kit of the present invention can be used for the prevention or treatment of liver disease.
본 발명에서 용어 "예방"은 (a) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 (b) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 결합체 (3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제를 포함하는 조합물 또는 조성물의 투여로 목적하는 질환, 예컨대 간 질환의 발병을 억제 또는 지연시키는 모든 행위를 의미하며, "치료"는 (a) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 (b) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 결합체 (3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제를 포함하는 조합물 또는 조성물의 투여로 목적하는 질환, 예컨대 간 질환의 증세가 호전되거나 이롭게 되는 모든 행위를 의미한다.In the present invention, the term "prevention" refers to (a) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate) and an FXR agonist; or (b) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate), a combination comprising an FXR agonist and an ACC inhibitor; or It refers to any action that inhibits or delays the onset of a desired disease, such as liver disease, by administration of the composition, and "treatment" refers to (a) a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (3 heavy agonist) or a conjugate thereof (a triple acting long-acting conjugate) and an FXR agonist; or (b) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate), a combination comprising an FXR agonist and an ACC inhibitor; or It refers to any action in which the symptoms of a target disease, such as liver disease, are improved or beneficial by administration of the composition.
본 발명에서 용어 "투여"는 어떠한 적절한 방법으로 환자에게 소정의 물질을 도입하는 것을 의미하며, 상기 조성물의 투여 경로는 특별히 이에 제한되지 않으나, 상기 조성물이 생체 내 표적에 도달할 수 있는 어떠한 일반적인 경로를 통하여 투여될 수 있으며, 예를 들어 복강 내 투여, 정맥 내 투여, 근육 내 투여, 피하 투여, 피내 투여, 경구 투여, 국소 투여, 비 내 투여, 폐 내 투여, 직장 내 투여 등이 될 수 있다.In the present invention, the term "administration" means introducing a predetermined substance to a patient by any suitable method, and the route of administration of the composition is not particularly limited thereto, but any general route through which the composition can reach an in vivo target It can be administered through, for example, intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intradermal administration, oral administration, topical administration, intranasal administration, intrapulmonary administration, rectal administration, etc. can be .
상기 (a) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 결합체 (3중 작용성 지속형 결합체) 및 FXR 작용제; 또는 (b) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 결합체 (3중 작용성 지속형 결합체), FXR 작용제 및 ACC 억제제는 병용으로 이용되어 간 질환의 예방 또는 치료에 사용될 수 있다.(a) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate) and an FXR agonist; or (b) a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a conjugate thereof (triple action long-acting conjugate), an FXR agonist and an ACC inhibitor are used in combination to liver It can be used for the prevention or treatment of diseases.
본 발명에서 용어 “간 질환”은 간에서 발병하는 질환을 의미하는 것으로, 대사성 간질환 또는 간 염증을 포함할 수 있으나, 이에 제한되는 것은 아니다. 상기 간질환의 대표적인 예로는, 비알코올성 지방간 질환(non-alcoholic fatty liver disease) 또는 담즙정체성 간질환(cholestasis liver disease) 등을 들 수 있다. 본 발명에 따른 상기 병용 요법에 사용되는 약학적 조성물, 조합물, 키트 등에 의해 그 질환이 예방 또는 치료될 수 있는 한, 간의 조직 및 기능에 이상이 생기는 다양한 간 질환은 본 발명의 간질환의 범주에 들어갈 수 있으나, 이에 제한되지 않는다.As used herein, the term “liver disease” refers to a disease occurring in the liver, and may include metabolic liver disease or liver inflammation, but is not limited thereto. Representative examples of the liver disease include non-alcoholic fatty liver disease or cholestasis liver disease. As long as the disease can be prevented or treated by the pharmaceutical composition, combination, kit, etc. used for the combination therapy according to the present invention, various liver diseases that cause abnormalities in the tissues and functions of the liver are within the scope of the liver disease of the present invention. can be entered, but is not limited thereto.
본 발명에서 용어 "비알코올성 지방간 질환"은 알코올 섭취 과거력이 없거나 알코올 섭취와 관련이 없으면서도 지방간을 동반하는 경우를 말한다. 지방간은 중성지방이 정상적인 경우와는 다르게 간 세포 내에 비정상적으로 침착되어 보이는 현상이 나타난 것을 말한다. 정상 간은 약 5%가 지방조직으로 구성되어 있으며 중성지방, 지방산, 인지질, 콜레스테롤 및 콜레스테롤 에스터가 지방의 주성분이나, 일단 지방간이 발생되면 대부분의 성분이 중성지방으로 대체되며, 중성지방의 양이 간 중량의 5%이상이면 지방간으로 진단된다. 지방간은 간세포 내의 지방대사 장애나 과잉지방을 운반하는 과정에서의 결함 등에 의하여 초래되는 것으로서, 주로 간에서의 지방대사 장애로 인하여 발생한다. 상기 지방간에서 축적된 지방의 대부분은 중성지방 (triglyceride)일 수 있다. 본 발명에 따른 조성물은 NAS (NAFLD activity score) 감소 및 섬유화를 억제하는 것을 확인한 바, 상기 비알코올성 지방간 질환에 포함되는 다양한 질환의 병변을 예방 또는 치료할 수 있음을 알 수 있다.In the present invention, the term "nonalcoholic fatty liver disease" refers to a case in which there is no history of alcohol intake or is accompanied by fatty liver although it is not related to alcohol intake. Fatty liver refers to a phenomenon in which triglycerides are abnormally deposited in liver cells, unlike normal cases. About 5% of a normal liver is composed of adipose tissue, and triglycerides, fatty acids, phospholipids, cholesterol, and cholesterol esters are the main components of fat. If it is more than 5% of the liver weight, it is diagnosed as fatty liver. Fatty liver is caused by disorders in fat metabolism in hepatocytes or defects in the process of transporting excess fat, and mainly occurs due to disorders in fat metabolism in the liver. Most of the fat accumulated in the fatty liver may be triglyceride. As it was confirmed that the composition according to the present invention inhibits NAS (NAFLD activity score) reduction and fibrosis, it can be seen that lesions of various diseases included in the nonalcoholic fatty liver disease can be prevented or treated.
상기 비알코올성 지방간질환은 단순 지방증, 간 염증, 비알코올성 지방간, 비알코올성 지방간염, 간경변, 간 섬유증, 간 부전 또는 간암 등이 이에 속하나, 본 발명의 조성물로 예방 또는 치료되는 비알코올성 지방간 질환은 제한 없이 포함된다.The nonalcoholic fatty liver disease includes simple steatosis, liver inflammation, nonalcoholic fatty liver, nonalcoholic steatohepatitis, cirrhosis, liver fibrosis, liver failure or liver cancer. included without
또 하나의 예로서, 상기 비알코올성 지방간 질환은 간 염증, 비알코올성 지방간염과 간 섬유증으로 이루어진 군에서 선택하는 적어도 하나의 질환일 수 있으나, 본 발명의 조성물로 예방 또는 치료되는 비알코올성 지방간 질환은 제한 없이 포함된다.As another example, the nonalcoholic fatty liver disease may be at least one disease selected from the group consisting of liver inflammation, nonalcoholic steatohepatitis and liver fibrosis, but the nonalcoholic fatty liver disease prevented or treated with the composition of the present invention is included without limitation.
본 발명에서, "비알코올성 지방간염(nonalcoholic steatohepatitis)”은 비알코올성 지방간질환의 하나로, 간 세포 괴사, 염증, 및 섬유화를 수반하는 간 질환의 대표적 예이다. 본 발명에 따른 조성물은 NAS (NAFLD activity score) 감소 및 섬유화를 억제하여 비알코올성 지방간염에 대한 효과를 나타낼 수 있으며, 구체적으로, 지방간, 간섬유증 또는 간경변을 수반한 비알코올성 지방간염; 또는 비알코올성 지방간염에 의한 간암에 대한 효과를 나타낼 수 있으나, 이에 제한되는 것은 아니다.In the present invention, "nonalcoholic steatohepatitis" is one of nonalcoholic fatty liver disease, and is a representative example of liver disease accompanied by liver cell necrosis, inflammation, and fibrosis. The composition according to the present invention contains NAS (NAFLD activity) score) reduction and suppression of fibrosis to exhibit an effect on nonalcoholic steatohepatitis, specifically, nonalcoholic steatohepatitis with fatty liver, liver fibrosis or cirrhosis; or liver cancer caused by nonalcoholic steatohepatitis However, the present invention is not limited thereto.
본 발명에서, “간 염증(liver inflamation)”은 간질환의 가장 큰 원인으로 간에 염증을 일으키는 질병을 의미하며, 원인과 증상에 따라서 급성간염과 만성간염으로 구별된다. 바이러스, 알코올, 약물, 면역 이상, 대사 질환 등을 주원인으로 한다. In the present invention, “liver inflamation” refers to a disease that causes inflammation of the liver as the biggest cause of liver disease, and is divided into acute hepatitis and chronic hepatitis according to the cause and symptoms. Viruses, alcohol, drugs, immune abnormalities, and metabolic diseases are the main causes.
본 발명에서, “간 섬유증 (liver fibrosis)”은 반복적인 간 손상에 대한 상처 회복 과정의 결과로, 간경변증과는 달리 가역적이고, 얇은 원섬유(thin fibril)로 구성되며, 결절(nodule) 형성이 없는 것으로 알려져 있고, 간 손상의 원인이 소실되면 정상회복이 가능할 수 있으나, 이러한 간 섬유증 과정이 반복적으로 지속되면 ECM(extra cellular matrix) 간의 교차결합(crosslinking)이 증가하여 결절(nodule)이 있는 비가역적인 간경변증으로 진행된다.In the present invention, "liver fibrosis" is a result of the wound healing process for repeated liver damage, and unlike liver cirrhosis, it is reversible, consists of thin fibrils, Normal recovery may be possible when the cause of liver damage is eliminated. progresses to cirrhosis of the liver.
본 발명의 조성물은 상기 약학적 조성물은 투여시 투여받은 개체에서 하이드록시프롤린 함량을 감소시킴으로써 간섬유증에 대한 예방 또는 치료 효과를 나타낼 수 있으나, 이에 제한되는 것은 아니다. 본 발명에 따른 결합체는 간의 섬유화 자체를 완화하는 효과를 가질 뿐만 아니라, 간의 섬유화를 수반하거나, 이에 기인하여 발병한 질환에도 효과를 나타낼 수 있다.The composition of the present invention may exhibit a preventive or therapeutic effect on liver fibrosis by reducing the hydroxyproline content in the subject to which the pharmaceutical composition is administered, but is not limited thereto. The conjugate according to the present invention not only has the effect of alleviating the fibrosis of the liver itself, but also can exhibit an effect on diseases accompanying or caused by fibrosis of the liver.
본 발명의 “간경변(liver cirrhosis)”은 간세포 재생 및 섬유조직의 증가를 반복하면서 발병하고 병리학적으로 괴사(necrosis), 염증(inflammation) 및 섬유화(fibrosis)가 수반되는 만성질환이며 궁극적으로 간 부전과 같은 간경변 합병증 및 간암 등의 질환으로 진행되어 사망에 이르게 한다. 특히, 초기에 자각증상이 없어 상당히 진행되어서야 발견되기 때문에, 간경변 등으로 진화되기 전의 상태인 간섬유증을 신속히 치료하는 것이 요구된다."liver cirrhosis" of the present invention is a chronic disease that occurs while repeating hepatocyte regeneration and increase in fibrous tissue and is pathologically accompanied by necrosis, inflammation and fibrosis, and ultimately liver failure It progresses to liver cirrhosis complications and diseases such as liver cancer, leading to death. In particular, since there are no subjective symptoms in the early stage and it is discovered only at a fairly advanced stage, it is required to rapidly treat liver fibrosis, which is a condition before it evolves into cirrhosis or the like.
본 발명의 “간 부전(liver decompensation)”은 바이러스성 간염, 간경변증, 약물 또는 알코올과 같은 간 손상 또는 간 질환으로 인해, 간 기능이 약화되고 간이 정상적인 생리 작용으로서의 단백질 합성과 대사 기능을 수행할 수 없는 상태를 의미한다. 진행 속도에 따라 급성 간부전 또는 만성 간부전으로 나뉘며, 다양한 합병증을 일으키는 것으로 알려져 있다. The "liver decompensation" of the present invention is due to liver damage or liver disease such as viral hepatitis, cirrhosis, drugs or alcohol, liver function is weakened, and the liver can perform protein synthesis and metabolic functions as normal physiological functions. means no state. It is divided into acute liver failure or chronic liver failure according to the rate of progression, and is known to cause various complications.
본 발명의 “간암(hepatocellular carcinoma)”은 간 세포에서 기원하는 악성 종양을 의미하며, 간세포 자체로부터 발생한 원발성 간암 (간세포암; hepatocellular carcinoma)과 다른 조직의 암이 간으로 전이되어 온 전이성 간암으로 구분할 수 있는데, 간암의 약 90% 이상은 원발성 간암이다. 주된 원인으로는 간염, 만성 간질환 외에 알코올, 흡연, 비만 등이 알려져 있다."Hepatocellular carcinoma" of the present invention means a malignant tumor originating from liver cells, and can be divided into primary liver cancer (hepatocellular carcinoma) arising from hepatocytes itself and metastatic liver cancer in which cancers of other tissues have metastasized to the liver. More than 90% of liver cancers are primary liver cancers. In addition to hepatitis and chronic liver disease, the main causes are alcohol, smoking, and obesity.
본 발명에서, “담즙정체증 (cholestasis)”은, 간에서 십이지장으로의 담즙의 유동이 느려지거나 차단된 병태이고, “담즙정체성 간질환(cholestasis liver disease)”은 간 내부에서, 담즙 형성이 각종 질환, 확장된 경정맥 영양, 또는 특정 약물 (예컨대, 일부 항생제) 의 부작용과 같은과 병태에 의해 방해된 것을 의미한다. 담즙정체증의 통상의 징후에는, 피로, 소양감 (가려움증), 황달, 및 황색종 (피하에의 고콜레스테롤 (cholesterol-rich) 물질의 증착) 이 포함된다. 담즙정체증의 영향은 극심하며 광범위하고, 이는 간 질환의 전신적 질환으로의 악화, 간부전, 및 간 이식의 필요를 초래한다. 담즙정체성 간질환의 원인으로는 급성 간염, 담관의 염증 등을 들 수 있다.In the present invention, "cholestasis" is a condition in which the flow of bile from the liver to the duodenum is slowed or blocked, and "cholestasis liver disease" is It is meant to be hampered by conditions such as disease, extended jugular vein nutrition, or side effects of certain medications (eg, some antibiotics). Common signs of cholestasis include fatigue, pruritus (itching), jaundice, and xanthomas (deposition of high cholesterol-rich substances under the skin). The effects of cholestasis are severe and widespread, leading to exacerbation of liver disease into systemic disease, liver failure, and the need for liver transplantation. Causes of cholestatic liver disease include acute hepatitis and inflammation of the bile ducts.
상기 담즙정체성 간질환에는, 원발성 담즙성 담관염 (PBC), 원발성 경화성 담관염 (PSC), 진행성 가족성 간내 담즙정체증 (PFIC), 및 알라질 증후군 (Alagille syndrome) (AS) 등이 포함될 수 있으나, 이에 제한되지 않는다.The cholestatic liver disease may include primary biliary cholangitis (PBC), primary sclerosing cholangitis (PSC), progressive familial intrahepatic cholestasis (PFIC), and Alagille syndrome (AS), etc. It is not limited thereto.
원발성 담즙성 담관염 (primary biliary cholangitis: PBC)으로도 알려진 원발성 담즙성 경변증은 원인 불명의 만성 담즙정체성 간 질환이다. 문맥 (portal) 및 문맥주위 (periportal) 염증으로 인한 진행성 담관 손상은 진행성 섬유증 및 궁극적 간경화를 일으킬 수 있다. 지금까지는 면역학적, 유전적 및 환경적 요인이 질환에 잠재적인 원인으로 알려져 있다. 원발성 담즙성 경변증은 주로 중년 여성에서 많이 나타나며, 증상은 초기 발현에서 피로, 가려움증 또는 미규명된 고지혈증이 또한 원발성 담즙성 경변증의 증상으로 나타날 수 있다. Primary biliary cirrhosis, also known as primary biliary cholangitis (PBC), is a chronic cholestatic liver disease of unknown etiology. Progressive bile duct damage due to portal and periportal inflammation can lead to progressive fibrosis and eventual cirrhosis of the liver. So far, immunological, genetic and environmental factors are known as potential causes of the disease. Primary biliary cirrhosis is mainly seen in middle-aged women, and the symptoms are fatigue, itchiness, or unexplained hyperlipidemia at the initial onset, and may also appear as symptoms of primary biliary cirrhosis.
현재까지는 원발성 담즙성 경화증이 면역-매개된 질환이라고 알려져 있으며, 구체적으로, 문맥 및 문맥주위 부위에서 T 림프구의 면역조직화학적 염색은 CD4-양성 및 CD8-음성 T 세포를 나타낸다. 또한, 이환된 개체의 무증상 제1-등급 동류 (relatives)에서 비정상 억제자 T-세포 활성이 보고되었다. 인터류킨이 변경된 면역 기능 및 섬유증에 기여함으로써 PBC의 발병기전에 역할을 할 수 있다는 것이 보고되었다 (G.J. Webb et al, J. Autoimmunity, 2015 Nov; 64: 42-52). To date, primary biliary sclerosis is known to be an immune-mediated disease, and specifically, immunohistochemical staining of T lymphocytes in the portal vein and periportal sites reveals CD4-positive and CD8-negative T cells. In addition, abnormal suppressor T-cell activity has been reported in asymptomatic Grade 1-relatives of affected individuals. It has been reported that interleukins may play a role in the pathogenesis of PBC by contributing to altered immune function and fibrosis (G.J. Webb et al, J. Autoimmunity, 2015 Nov; 64: 42-52).
PBC의 치료방법은 우르소데옥시콜산 (UDSA) 및 오베티콜산 (OCA)을 이용한 담즙산 치료법이다. PBC에서 두 약물의 작용 메커니즘은 FXR 및 TGFR-5를 활성화시켜 항-염증성 효과를 발휘하게 하는 이들의 능력과 연관된다. 그러나, UDCA로 치료된 환자의 약 40%에서 충분한 생화학적 반응을 달성되지 않았다. The treatment method for PBC is bile acid therapy using ursodeoxycholic acid (UDSA) and obeticholic acid (OCA). The mechanism of action of both drugs in PBC is related to their ability to activate FXR and TGFR-5 to exert anti-inflammatory effects. However, sufficient biochemical responses were not achieved in about 40% of patients treated with UDCA.
원발성 경화성 담관염(Primary sclerosing cholangitis: PSC)는 원인 모를 간내/간외 담도의 염증과 섬유화로 인해 발생되는 만성 진행성 담즙 정체성 간질환이다. 구체적으로, 담관 및 담도의 염증성 질환으로서 질환이 진행되면 섬유화가 일어나 담관벽이 두꺼워지면서 좁아지거나 협착이 오는 질환이다. 아직까지 원인은 불명이나, 유전적 요인, 환경적 요인, 이와 관련된 면역반응 등 다양한 인자가 복합적으로 원인으로 추정하고 있다. Primary sclerosing cholangitis (PSC) is a chronic progressive cholestatic liver disease caused by inflammation and fibrosis of the intrahepatic/extrahepatic biliary tract of unknown etiology. Specifically, as an inflammatory disease of the bile duct and biliary tract, as the disease progresses, fibrosis occurs and the bile duct wall thickens and narrows or narrows. The cause is still unknown, but various factors such as genetic factors, environmental factors, and related immune responses are assumed to be the cause of the complex.
혈액을 통한 간 기능 검사에서 alkaline phosphatase 수치 상승, aminotransferase 수치 상승, 감마글로불린혈증 등이 보이면 원발성 경화성 담관염으로 진단하고 있다. If liver function tests using blood show elevated alkaline phosphatase levels, elevated aminotransferase levels, and gamma globulinemia, the diagnosis of primary sclerosing cholangitis is indicated.
PSC의 치료방법은 아직까지 명확하게 보고된 바 없으며, 간이식 수술이 근본적으로 치료할 수 있는 유일한 치료방법이다. The treatment method for PSC has not yet been clearly reported, and liver transplantation is the only treatment method that can fundamentally treat it.
이에, 환자 편의성 확보 및 부작용 없이 PBS, 및 PSC를 치료할 수 있는 약물의 개발이 여전히 요구되고 있으며, 본 발명에 따른 3중 작용제 또는 이의 지속형 결합체 및 FXR 작용제와의 병용; 또는 3중 작용제 또는 이의 지속형 결합체, FXR 작용제 및 ACC 억제제와의 병용 요법은 상기 질환을 치료하기에 적합하다.Accordingly, there is still a need to develop a drug capable of treating PBS and PSC without securing patient convenience and side effects, and the triple agent or long-acting conjugate thereof according to the present invention and combination with an FXR agent; or combination therapy with a triple agonist or a long-acting conjugate thereof, an FXR agonist and an ACC inhibitor is suitable for treating said disease.
본 발명에 따른 조성물은 기존 비알코올성 지방간질환 치료제의 부작용인 체중 증가가 없거나 그 정도가 상대적으로 낮은 것을 특징으로 할 수 있다.The composition according to the present invention may be characterized in that there is no or a relatively low degree of weight gain, which is a side effect of the existing therapeutic agent for nonalcoholic fatty liver disease.
또한, 상기 조성물은 (a) 섬유증 마커 (fibrosis marker)인 콜라겐-1a의 발현 또는 활성 감소; (b) 염증 유발 마커 (pro-inflammatory marker)인 TNF-alpha (tumor necrosis factor-α)의 발현 또는 활성 감소; (c) 지방 생성 마커 (lipogenesis marker)인 SREBP-1c (sterol regulatory element binding protein-1c)의 발현 또는 활성 감소; (d) 간 내 중성 지방 감소; 및 (e) 혈액 내 콜레스테롤 감소와 같은 (a) 내지 (e)의 특성 중 하나 이상의 특성을 수행하여, 간 질환을 예방 또는 치료할 수 있다. 그러나, 이에 제한되는 것은 아니다. In addition, the composition may (a) decrease the expression or activity of collagen-1a, a fibrosis marker; (b) decreased expression or activity of TNF-alpha (tumor necrosis factor-α), a pro-inflammatory marker; (c) decreased expression or activity of sterol regulatory element binding protein-1c (SREBP-1c), a lipogenesis marker; (d) reduction of triglycerides in the liver; and (e) one or more of the properties of (a) to (e), such as reducing blood cholesterol, to prevent or treat liver disease. However, the present invention is not limited thereto.
또한 상기 조성물은 NAS (NAFLD activity score)를 감소시켜 간 질환, 그 예로 비알코올성 지방간 질환을 예방 또는 치료할 수 있다.In addition, the composition can reduce NAS (NAFLD activity score) to prevent or treat liver disease, for example, non-alcoholic fatty liver disease.
또한 상기 조성물은 간 조직내 섬유증 지표로 알려진 하이드록시프롤린 (hydroxyprolin)을 감소시켜 간 질환, 그 예로 비알코올성 지방간 질환을 예방 또는 치료할 수 있다.In addition, the composition can prevent or treat liver disease, for example, non-alcoholic fatty liver disease by reducing hydroxyprolin, which is known as an indicator of fibrosis in liver tissue.
본 발명의 약학적 조성물은 약학적으로 허용가능한 담체, 부형제 또는 희석제를 추가로 포함할 수 있다. 이러한 약학적으로 허용가능한 담체, 부형제, 또는 희석제는 비자연적으로 발생된 것일 수 있다. The pharmaceutical composition of the present invention may further include a pharmaceutically acceptable carrier, excipient or diluent. Such pharmaceutically acceptable carriers, excipients, or diluents may be non-naturally occurring.
본 발명에서 용어 "약학적으로 허용가능한"이란 치료효과를 나타낼 수 있을 정도의 충분한 양과 부작용을 일으키지 않는 것을 의미하며, 질환의 종류, 환자의 연령, 체중, 건강, 성별, 환자의 약물에 대한 민감도, 투여 경로, 투여 방법, 투여횟수, 치료 기간, 배합 또는 동시 사용되는 약물 등 의학 분야에 잘 알려진 요소에 따라 당업자에 의해 용이하게 결정될 수 있다.In the present invention, the term "pharmaceutically acceptable" means a sufficient amount to exhibit a therapeutic effect and does not cause side effects, and the type of disease, the patient's age, weight, health, sex, and the patient's sensitivity to the drug , administration route, administration method, frequency of administration, treatment period, combination or drugs used at the same time can be easily determined by those skilled in the art according to factors well known in the medical field.
본 발명의 펩타이드를 포함한 약학적 조성물은 약학적으로 허용가능한 부형제를 추가로 포함할 수 있다. 상기 부형제는 특별히 이에 제한되지는 않으나, 경구 투여 시에는 결합제, 활택제, 붕해제, 가용화제, 분산제, 안정화제, 현탁화제, 색소, 향료 등을 사용할 수 있고, 주사제의 경우에는 완충제, 보존제, 무통화제, 가용화제, 등장화제, 안정화제 등을 혼합하여 사용할 수 있으며, 국소투여용의 경우에는 기제, 부형제, 윤활제, 보존제 등을 사용할 수 있다. The pharmaceutical composition including the peptide of the present invention may further include a pharmaceutically acceptable excipient. The excipient is not particularly limited thereto, but in the case of oral administration, a binder, a lubricant, a disintegrant, a solubilizer, a dispersing agent, a stabilizer, a suspending agent, a dye, a flavoring agent, etc. may be used, and in the case of an injection, a buffer, a preservative, An analgesic agent, a solubilizer, an isotonic agent, a stabilizer, etc. can be mixed and used, and in the case of topical administration, a base, excipient, lubricant, preservative, etc. can be used.
본 발명의 조성물의 제형은 상술한 바와 같은 약학적으로 허용가능한 부형제와 혼합하여 다양하게 제조될 수 있다. 예를 들어, 경구 투여 시에는 정제, 트로키, 캡슐, 엘릭서, 서스펜션, 시럽, 웨이퍼 등의 형태로 제조할 수 있으며, 주사제의 경우에는 단위 투약 앰플 또는 다수회 투약 형태로 제조할 수 있다. 기타, 용액, 현탁액, 정제, 환약, 캡슐, 서방형 제제 등으로 제형화할 수 있다.The formulation of the composition of the present invention can be prepared in various ways by mixing with the pharmaceutically acceptable excipients as described above. For example, in the case of oral administration, it may be prepared in the form of tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like, and in the case of injections, it may be prepared in the form of unit dose ampoules or multiple doses. In addition, it can be formulated as a solution, suspension, tablet, pill, capsule, sustained release formulation, and the like.
한편, 제제화에 적합한 담체, 부형제 및 희석제의 예로는 락토스, 덱스트로스, 수크로스, 솔비톨, 만니톨, 자일리톨, 에리스리톨, 말티톨, 전분, 아카시아, 알지네이트, 젤라틴, 칼슘 포스페이트, 칼슘 실리케이트, 셀룰로스, 메틸 셀룰로스, 미정질 셀룰로스, 폴리비닐피롤리돈, 물, 메틸하이드록시벤조에이트, 프로필하이드록시벤조에이트, 탈크, 마그네슘 스테아레이트 또는 광물유 등이 사용될 수 있다. 또한, 충진제, 항응집제, 윤활제, 습윤제, 향료, 방부제 등을 추가로 포함할 수 있다. Meanwhile, examples of suitable carriers, excipients and diluents for formulation include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, Microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, or mineral oil may be used. In addition, it may further include a filler, an anti-aggregating agent, a lubricant, a wetting agent, a flavoring agent, a preservative, and the like.
또한, 본 발명의 약학적 조성물은 정제, 환제, 산제, 과립제, 캡슐제, 현탁제, 내용액제, 유제, 시럽제, 멸균된 수용액, 비수성용제, 동결건조제제 및 좌제로 이루어진 군으로부터 선택되는 어느 하나의 제형을 가질 수 있다.In addition, the pharmaceutical composition of the present invention is any one selected from the group consisting of tablets, pills, powders, granules, capsules, suspensions, internal solutions, emulsions, syrups, sterile aqueous solutions, non-aqueous solvents, freeze-dried preparations and suppositories may have the form of
또한, 상기 조성물은 약학적 분야에서 통상의 방법에 따라 환자의 신체 내 투여에 적합한 단위 투여형의 제제, 구체적으로는 단백질 의약품의 투여에 유용한 제제 형태로 제형화시켜 당업계에서 통상적으로 사용하는 투여 방법을 이용하여 경구, 또는 피부, 정맥 내, 근육 내, 동맥 내, 골수 내, 수막강 내, 심실 내, 폐, 경피, 피하, 복 내, 비강 내, 소화관 내, 국소, 설하, 질 내 또는 직장 경로를 포함하는 비경구 투여 경로에 의하여 투여될 수 있으나, 이들에 한정되는 것은 아니다.In addition, the composition is formulated in a dosage form suitable for administration in a patient's body according to a conventional method in the pharmaceutical field, specifically, a formulation useful for administration of a protein drug, and administration commonly used in the art. Oral, or dermal, intravenous, intramuscular, intraarterial, intramedullary, intrathecal, intraventricular, pulmonary, transdermal, subcutaneous, intraperitoneal, intranasal, gastrointestinal, topical, sublingual, intravaginal or It may be administered by parenteral routes of administration including, but not limited to, rectal routes.
또한, 상기 3중 작용제 또는 이의 지속형 결합체는 생리식염수 또는 유기용매와 같이 약제로 허용된 여러 전달체(carrier)와 혼합하여 사용될 수 있고, 안정성이나 흡수성을 증가시키기 위하여 글루코스, 수크로스 또는 덱스트란과 같은 탄수화물, 아스코르브산(ascorbic acid) 또는 글루타티온과 같은 항산화제(antioxidants), 킬레이트제, 저분자 단백질 또는 다른 안정화제(stabilizers) 등이 약제로 사용될 수 있다.In addition, the triple agonist or its long-acting conjugate can be used by mixing with various pharmaceutically acceptable carriers such as physiological saline or organic solvents, and with glucose, sucrose or dextran to increase stability or absorption. Carbohydrates, antioxidants such as ascorbic acid or glutathione, chelating agents, low molecular weight proteins or other stabilizers may be used as agents.
본 발명의 약학적 조성물의 투여량과 횟수는 치료할 질환, 투여 경로, 환자의 연령, 성별 및 체중 및 질환의 중등도 등의 여러 관련 인자와 함께, 활성성분인 약물의 종류에 따라 결정된다. . 구체적으로, 본 발명의 조성물은 상기 펩타이드 또는 이를 포함하는 지속형 결합체를 약학적 유효량으로 포함하는 것일 수 있으나, 이에 제한되지 않는다.The dosage and frequency of administration of the pharmaceutical composition of the present invention is determined according to the type of drug as the active ingredient, together with several related factors such as the disease to be treated, the route of administration, the age, sex and weight of the patient, and the severity of the disease. . Specifically, the composition of the present invention may include the peptide or a long-acting conjugate comprising the same in a pharmaceutically effective amount, but is not limited thereto.
상기 펩타이드 또는 지속형 결합체를 약학적 유효량으로 포함하는 것은, 펩타이드 또는 지속형 결합체로 인한 목적하는 약리 활성(예를 들어, 간 질환의 예방, 개선 또는 치료)을 얻을 수 있는 정도를 의미하고, 또한 투여되는 개체에서 독성 또는 부작용이 일어나지 않거나 미미한 수준으로서 약학적으로 허용되는 수준을 의미할 수 있으나, 이에 제한되지 않는다. 이와 같은 약학적 유효량은 투여 횟수, 환자, 제형 등을 종합적으로 고려하여 결정될 수 있다.The inclusion of the peptide or the long-acting conjugate in a pharmaceutically effective amount means the degree to which the desired pharmacological activity (eg, prevention, improvement or treatment of liver disease) can be obtained due to the peptide or the long-acting conjugate, and also It may mean a pharmaceutically acceptable level as a level that does not cause toxicity or side effects in the administered subject or is insignificant, but is not limited thereto. Such a pharmaceutically effective amount may be determined by comprehensively considering the number of administration, patient, formulation, and the like.
특별히 이에 제한되지 않으나, 본 발명의 상기 약학적 조성물은 상기 성분 (유효성분)을 0.01 내지 99% 중량 대 부피로 함유할 수 있다. Although not particularly limited thereto, the pharmaceutical composition of the present invention may contain the component (active ingredient) in an amount of 0.01 to 99% by weight to volume.
본 발명의 조성물의 총 유효량은 단일 투여량 (single dose)으로 환자에게 투여될 수 있으며, 다중 투여량 (multiple dose)으로 장기간 투여되는 분할 치료 방법 (fractionated treatment protocol)에 의해 투여될 수 있다. 본 발명의 약학적 조성물은 질환의 정도에 따라 유효성분의 함량을 달리할 수 있다. 구체적으로, 본 발명의 펩타이드 또는 결합체의 바람직한 전체 용량은 하루에 환자 체중 1 kg당 약 0.0001 mg 내지 500 mg일 수 있으나, 이에 제한되지 않는다. 그러나 상기 결합체의 용량은 약학적 조성물의 투여 경로 및 치료 횟수뿐만 아니라 환자의 연령, 체중, 건강 상태, 성별, 질환의 중증도, 식이 및 배설율 등 다양한 요인들을 고려하여 환자에 대한 유효 투여량이 결정되는 것이므로, 이러한 점을 고려할 때 당분야의 통상적인 지식을 가진 자라면 상기 본 발명의 조성물의 특정한 용도에 따른 적절한 유효 투여량을 결정할 수 있을 것이다. 본 발명에 따른 약학적 조성물은 본 발명의 효과를 보이는 한 그 제형, 투여 경로 및 투여 방법에 특별히 제한되지 아니한다.The total effective amount of the composition of the present invention may be administered to a patient as a single dose, or may be administered by a fractionated treatment protocol in which multiple doses are administered for a long period of time. The pharmaceutical composition of the present invention may vary the content of the active ingredient according to the severity of the disease. Specifically, the preferred total dose of the peptide or conjugate of the present invention may be about 0.0001 mg to 500 mg per 1 kg of the patient's body weight per day, but is not limited thereto. However, the dosage of the conjugate is determined by considering various factors such as the age, weight, health status, sex, severity of disease, diet and excretion rate of the patient, as well as the administration route and number of treatments of the pharmaceutical composition, and the effective dosage for the patient is determined. Therefore, in consideration of this point, those of ordinary skill in the art will be able to determine an appropriate effective dosage according to the specific use of the composition of the present invention. The pharmaceutical composition according to the present invention is not particularly limited in its formulation, administration route and administration method as long as the effect of the present invention is exhibited.
본 발명의 3중 작용제 또는 이의 지속형 결합체는 FXR 작용제; 또는 FXR 작용제 및 ACC 억제제와 적절한 비율로 병용 투여될 수 있다. 예를 들어, 1회 투여량 기준으로 본 발명의 3중 작용제 또는 이의 지속형 결합체는 약 0.1 내지 약 100 mg, 약 0.5 내지 약 80 mg, 약 1 내지 약 60 mg, 약 1 내지 약 25 mg, 약 5 내지 약 20 mg, 또는 약 10 내지 약 15 mg으로 투여되고, FXR 작용제는 약 1 내지 약 100 mg, 약 1 내지 약 250 mg, 약 5 내지 약 250 mg, 약 5 내지 약 50 mg, 약 5 내지 약 40 mg, 약 5 내지 약 40 mg, 약 5 내지 약 25 mg, 약 5 내지 약 30 mg,약 90 내지 약 250 mg, 약 140 내지 약 250 mg,약 140 내지 약 200 mg으로 투여되고, ACC 억제제는 약 1 내지 1000 mg, 약 5 내지 약 1000 mg, 약 5 내지 150 mg, 약 5 내지 약 50 mg, 약 5 내지 약 30 mg, 약 5 내지 약 750 mg, 약 10 내지 약 550 mg으로 투여되는 것일 수 있으나, 이에 제한되지 않는다.The triple agonist or long-acting conjugate thereof of the present invention may be an FXR agonist; or an FXR agonist and an ACC inhibitor in an appropriate ratio. For example, on a single dose basis, the triple agent of the present invention or a long-acting conjugate thereof may be administered in an amount of about 0.1 to about 100 mg, about 0.5 to about 80 mg, about 1 to about 60 mg, about 1 to about 25 mg, administered at about 5 to about 20 mg, or about 10 to about 15 mg, and the FXR agonist is about 1 to about 100 mg, about 1 to about 250 mg, about 5 to about 250 mg, about 5 to about 50 mg, about 5 to about 40 mg, about 5 to about 40 mg, about 5 to about 25 mg, about 5 to about 30 mg, about 90 to about 250 mg, about 140 to about 250 mg, about 140 to about 200 mg, and , the ACC inhibitor is about 1 to 1000 mg, about 5 to about 1000 mg, about 5 to 150 mg, about 5 to about 50 mg, about 5 to about 30 mg, about 5 to about 750 mg, about 10 to about 550 mg It may be administered as, but is not limited thereto.
본 발명의 3중 작용제 또는 이의 지속형 결합체는 FXR 작용제; 또는 FXR 작용제 및 ACC 억제제와 적절한 비율로 병용 투여되어 우수한 간 질환 치료 효과를 나타낼 수 있다.The triple agonist or long-acting conjugate thereof of the present invention may be an FXR agonist; Alternatively, it may be administered in combination with an FXR agonist and an ACC inhibitor in an appropriate ratio to exhibit an excellent therapeutic effect on liver disease.
본 발명의 약학적 조성물은 생체 내 지속성 및 역가가 우수하여, 본 발명의 약학적 제제의 투여 횟수 및 빈도를 현저하게 감소시킬 수 있으나 이에 제한되지 않는.The pharmaceutical composition of the present invention has excellent in vivo persistence and potency, and can significantly reduce the number and frequency of administration of the pharmaceutical preparation of the present invention, but is not limited thereto.
본 발명을 구현하는 다른 하나의 양태는 FXR 작용제와 병용하는 것을 특징으로 하는 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체)를 포함하는, 간 질환의 예방 또는 치료용 약학적 조성물을 제공한다.Another embodiment embodying the present invention is a substance having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor (triple agonist) or a long-acting conjugate thereof (triple), characterized in that it is used in combination with an FXR agonist It provides a pharmaceutical composition for preventing or treating liver disease, including a long-acting long-acting conjugate).
또 하나의 구체예로서, 상기 약학적 조성물은 추가로 ACC 억제제와 병용하는 것을 특징으로 할 수 있다.In another embodiment, the pharmaceutical composition may be further characterized in that it is used in combination with an ACC inhibitor.
상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제, ACC 작용제를 포함하는 조성물, 예방, 치료, 간 질환 및 약학적 조성물에 대해서는 앞서 설명한 바와 같다.A substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist, a composition comprising an ACC agonist, prophylaxis, Treatment, liver disease and pharmaceutical compositions are the same as described above.
본 발명을 구현하는 다른 하나의 양태는 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), 및 (ii) FXR 작용제를 이를 필요로 하는 개체에 투여하는 단계를 포함하는, 간 질환의 예방 또는 치료 방법을 제공한다.Another aspect embodying the present invention is (i) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), and (ii) administering an FXR agonist to a subject in need thereof.
본 발명을 구현하는 다른 하나의 양태는, (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), (ii) FXR 작용제, 및 (iii) ACC 억제제를 이를 필요로 하는 개체에 투여하는 단계를 포함하는, 간 질환의 예방 또는 치료 방법일 수 있다.Another aspect embodying the present invention is (i) a substance having activity on a glucagon receptor, a GLP-1 receptor, and a GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate) , (ii) an FXR agonist, and (iii) an ACC inhibitor to a subject in need thereof.
본 발명을 구현하는 다른 하나의 양태는 간 질환의 치료가 필요한 개체에, 약학적 유효량으로 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체)를 투여하는 단계; 및 (ii) FXR 작용제를 투여하는 단계를 포함하며, 위 (i)과 (ii)의 단계는 순차적, 역순 또는 동시에 이루어지는, 간 질환의 예방 또는 치료 방법을 제공한다.Another aspect embodying the present invention is a pharmaceutically effective amount of (i) a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonist) or its administering a long-acting conjugate (triple-acting long-acting conjugate); and (ii) administering an FXR agonist, wherein the steps (i) and (ii) are sequential, reversed, or simultaneous.
본 발명을 구현하는 다른 하나의 양태는 간 질환의 치료가 필요한 개체에, 약학적 유효량으로 (i) 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체)를 투여하는 단계; (ii) FXR 작용제를 투여하는 단계; 및 (iii) ACC 억제제를 포함하며, 위 (i), (ii), (iii)의 단계는 순차적, 역순 또는 동시에 이루어지는, 간 질환의 예방 또는 치료 방법을 제공한다.Another aspect embodying the present invention is a pharmaceutically effective amount of (i) a substance having activity on glucagon receptors, GLP-1 receptors, and GIP receptors (triple agonist) or its administering a long-acting conjugate (triple-acting long-acting conjugate); (ii) administering an FXR agonist; and (iii) an ACC inhibitor, wherein the steps of (i), (ii), and (iii) are sequential, reversed, or simultaneous.
본 발명의 예방 또는 치료 방법의 각 약효 성분들을 투여하는 각각의 단계는 어느 순서로 이루어지든 무방하며, (i), (ii), (iii)은 순서를 의미하는 것이 아니며, 유효성분을 구분 짓는 것으로 각 단계는 순차적, 역순 또는 동시에 이루어질 수 있으나, 이에 제한되는 것은 아니다.Each step of administering each active ingredient of the prophylactic or therapeutic method of the present invention may be performed in any order, and (i), (ii), and (iii) do not mean the order, and Each step may be performed sequentially, in reverse order, or simultaneously, but is not limited thereto.
상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제, ACC 작용제를 포함하는 조성물, 예방, 치료, 간 질환 및 약학적 조성물에 대해서는 앞서 설명한 바와 같다.A substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist, a composition comprising an ACC agonist, prophylaxis, Treatment, liver disease and pharmaceutical compositions are the same as described above.
본 발명에서 상기 개체는 간 질환이 의심되는 개체로서, 상기 간 질환 의심 개체는 해당 질환이 발병하였거나 발병할 수 있는 인간을 포함한 쥐, 가축 등을 포함하는 포유 동물을 의미하나, 본 발명의 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체); 및 (a) FXR 작용제, 또는 (b) FXR 작용제 및 ACC 억제제를 포함하는 상기 조성물로 치료 가능한 개체는 제한 없이 포함된다. 또한, 본 발명의 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체); 및 (a) FXR 작용제, 또는 (b) FXR 작용제 및 ACC 억제제를 포함하는 약학적 조성물을 간 질환 의심 개체에 투여함으로써, 개체를 효율적으로 치료할 수 있다. 간 질환에 대해서는 상기에서 설명한 바와 같다.In the present invention, the subject is a subject suspected of liver disease, and the subject suspected of liver disease refers to mammals including mice, livestock, etc. including humans that have or may develop the disease, but the glucagon receptor of the present invention , a substance having activity on the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (a) an FXR agonist, or (b) an FXR agonist and an ACC inhibitor. In addition, substances having activity on the glucagon receptor, GLP-1 receptor, and GIP receptor of the present invention (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (a) an FXR agonist, or (b) an FXR agonist and an ACC inhibitor, by administering to the subject suspected of liver disease, the subject may be efficiently treated. Liver disease is the same as described above.
본 발명의 방법은 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체); 및 (a) FXR 작용제, 또는 (b) FXR 작용제 및 ACC 억제제를 포함하는 조합물 또는 약학적 조성물을 약학적 유효량으로 투여하는 것을 포함할 수 있다. 본 발명에 따른 방법은 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체); 및 (a) FXR 작용제, 또는 (b) FXR 작용제 및 ACC 억제제를 하나의 제제로서 투여하거나, 또는 개별 제제를 동시, 개별, 순차 또는 역순으로 투여하는 것일 수 있으나, 이에 제한되지 않는다.The method of the present invention comprises a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (a) an FXR agonist, or (b) a combination or pharmaceutical composition comprising an FXR agonist and an ACC inhibitor, in a pharmaceutically effective amount. The method according to the present invention comprises a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate); and (a) the FXR agonist, or (b) the FXR agonist and the ACC inhibitor are administered as one agent, or the separate agents are administered simultaneously, separately, sequentially or in reverse order.
본 발명의 용어, "투여"는 어떠한 적절한 방법으로 환자(개체)에게 소정의 물질을 도입하는 것을 의미한다. 상기 3중 작용제 또는 이의 지속형 결합체, FXR 작용제, ACC 억제제의 투여 경로는 특별히 이에 제한되지 않으나, 상기 3중 작용제 또는 이의 지속형 결합체, FXR 작용제, ACC 억제제가 생체 내 표적에 도달할 수 있는 어떠한 일반적인 경로를 통하여 투여될 수 있으며, 예를 들어 복강 내 투여, 정맥내 투여, 근육내 투여, 피하 투여, 피내 투여, 경구 투여, 국소 투여, 비내 투여, 폐내 투여, 직장 내 투여 등이 될 수 있다.As used herein, the term “administration” means introducing a given substance into a patient (individual) by any suitable method. The route of administration of the triple agonist or long-acting conjugate thereof, FXR agonist, or ACC inhibitor is not particularly limited thereto, but the triple agonist or long-acting conjugate thereof, FXR agonist, or any ACC inhibitor capable of reaching an in vivo target It may be administered through a general route, for example, intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intradermal administration, oral administration, topical administration, intranasal administration, intrapulmonary administration, rectal administration, etc. can be .
본 발명의 방법에서, 상기 3중 작용제 또는 이의 지속형 결합체, FXR 작용제, ACC 억제제는 동일한 투여 경로로 투여되거나, 또는 서로 다른 투여 경로로 투여될 수 있으며, 병용 투여되는 약물의 투여 경로는 서로 독립적일 수 있다.In the method of the present invention, the triple agonist or long-acting conjugate thereof, FXR agonist, and ACC inhibitor may be administered by the same administration route or may be administered by different administration routes, and the administration routes of the drugs administered in combination are independent of each other can be
이에 제한되지 않으나, 본 발명의 3중 작용제 또는 이의 지속형 결합체, FXR 작용제; 또는 3중 작용제 또는 이의 지속형 결합체, FXR 작용제, 및 ACC 억제제를 포함하는 조성물은 1일에 1회, 2일에 1회, 3일에 1회, 1주에 1회, 2주에 1회, 4주에 1회, 또는 1개월에 1회 투여될 수 있으나, 이에 제한되지 않는다.Without being limited thereto, the triple agonist of the present invention or a long-acting conjugate thereof, an FXR agonist; or a composition comprising a triple agonist or a long-acting combination thereof, an FXR agonist, and an ACC inhibitor once a day, once every 2 days, once every 3 days, once a week, once every two weeks , may be administered once every 4 weeks, or once a month, but is not limited thereto.
한편, FXR 작용제, 및 ACC 억제제는 본 발명의 3중 작용제 또는 이의 지속형 결합체 또는 이를 포함하는 조성물과 병용 투여됨으로써 간 질환 예방 또는 치료 효과를 발휘할 수 있는 한, 투여 빈도는 제한되지 않으나, 예를 들어, 1일에 1회, 2일에 1회, 3일에 1회, 1주에 1회, 2주에 1회, 4주에 1회, 또는 1개월에 1회 투여될 수 있다. 또한, 상기 3중 작용제 또는 이의 지속형 결합체와 동일한 간격으로 투여되거나, 또는 다른 간격, 예를 들어 1일, 2일, 3일, 4일, 5일, 6일, 7일, 8일, 9일, 또는 10일 이상의 간격을 두고 투여되는 것일 수 있으나, 이에 제한되는 것은 아니다,On the other hand, the FXR agonist and the ACC inhibitor are administered in combination with the triple agonist of the present invention, or a long-acting conjugate thereof, or a composition comprising the same, as long as it can exert a preventive or therapeutic effect on liver disease, but the frequency of administration is not limited, but for example For example, once a day, once every 2 days, once every 3 days, once a week, once every 2 weeks, once every 4 weeks, or once a month. In addition, the triple agent or long-acting combination thereof is administered at the same interval, or at a different interval, for example, 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days. It may be administered at intervals of one day, or 10 days or more, but is not limited thereto,
적합한 총 1일 사용량은 올바른 의학적 판단범위 내에서 처치의에 의해 결정될 수 있으며, 1회 또는 수회로 나누어 투여할 수 있다. 그러나 본 발명의 목적상, 특정 환자에 대한 구체적인 치료적 유효량은 달성하고자 하는 반응의 종류와 정도, 경우에 따라 다른 제제가 사용되는지의 여부를 비롯한 구체적 조성물, 환자의 연령, 체중, 일반 건강 상태, 성별 및 식이, 투여 시간, 투여 경로 및 조성물의 분비율, 치료기간, 구체적 조성물과 함께 사용되거나 동시 사용되는 약물을 비롯한 다양한 인자와 의약 분야에 잘 알려진 유사 인자에 따라 다르게 적용하는 것이 바람직하다.A suitable total daily amount may be determined by a treating physician within the scope of sound medical judgment, and may be administered once or divided into several doses. However, for the purposes of the present invention, a specific therapeutically effective amount for a particular patient depends on the type and extent of the response to be achieved, the specific composition, including whether other agents are used, if necessary, the specific composition, the patient's age, weight, general health, It is preferable to apply differently depending on various factors including sex and diet, administration time, administration route and secretion rate of the composition, treatment period, drugs used together or concurrently with a specific composition, and similar factors well known in the pharmaceutical field.
본 발명을 구현하는 또 하나의 양태는 상기 조합물, 상기 질환의 예방 또는 치료용 약학적 조성물, 또는 상기 약학적 키트를 이를 필요로 하는 개체에 투여 및/또는 사용하는 단계를 포함하는 간 질환의 예방 또는 치료 방법을 제공한다.Another aspect embodying the present invention is a liver disease comprising administering and/or using the combination, a pharmaceutical composition for the prevention or treatment of the disease, or the pharmaceutical kit to an individual in need thereof. A method of prevention or treatment is provided.
상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제, ACC 작용제를 포함하는 조합물, 키트, 조성물, 예방, 치료, 간 질환 및 약학적 조성물에 대해서는 앞서 설명한 바와 같다.A combination, kit comprising a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist, and an ACC agonist , composition, prevention, treatment, liver disease and pharmaceutical composition as described above.
본 발명을 구현하는 또 하나의 양태는 상기 조합물, 약학적 조성물, 또는 약학적 키트의 간 질환의 예방 또는 치료 용도 및/또는 간 질환의 예방 또는 치료를 위한 약제의 제조를 위한 용도를 제공한다.Another aspect embodying the present invention provides the use of the combination, pharmaceutical composition, or pharmaceutical kit for the prophylaxis or treatment of liver disease and/or the use of a medicament for the prophylaxis or treatment of liver disease. .
상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 물질 (3중 작용제) 또는 이의 지속형 결합체 (3중 작용성 지속형 결합체), FXR 작용제, ACC 작용제를 포함하는 조합물, 키트, 조성물, 예방, 치료, 간 질환 및 약학적 조성물에 대해서는 앞서 설명한 바와 같다.A combination, kit comprising a substance having activity on the glucagon receptor, the GLP-1 receptor, and the GIP receptor (triple agonist) or a long-acting conjugate thereof (triple-acting long-acting conjugate), an FXR agonist, and an ACC agonist , composition, prevention, treatment, liver disease and pharmaceutical composition as described above.
이하, 하기 실시예에 의하여 본 발명을 보다 상세하게 설명한다. 단, 하기 실시예는 본 발명을 예시하기 위한 것일 뿐 본 발명의 범위가 이들로 한정되는 것은 아니다.Hereinafter, the present invention will be described in more detail by way of Examples. However, the following examples are only for illustrating the present invention, and the scope of the present invention is not limited thereto.
실시예 1: 삼중 활성체의 제조Example 1: Preparation of triple activator
GLP-1 수용체, GIP 수용체 및 글루카곤 수용체에 모두 활성을 나타내는 삼중 활성체를 제조하여, 하기 표 1에 이의 서열을 나타냈다.A triple activator exhibiting activity on all GLP-1 receptors, GIP receptors and glucagon receptors was prepared, and its sequences are shown in Table 1 below.
서열번호SEQ ID NO: 서열order 정보information
1One H X Q G T F T S D V S S Y L D G Q A A K E F I A W L V K G C H X Q G T F T S D V S S Y L D G Q A A K E F I A W L V K G C 　
22 H X Q G T F T S D V S S Y L D G Q A Q K E F I A W L V K G C H X Q G T F T S D V S S Y L D G Q A Q K E F I A W L V K G C 　
33 H X Q G T F T S D V S S Y L L G Q A A K Q F I A W L V K G G G P S S G A P P P S C H X Q G T F T S D V S S Y L L G Q A A K Q F I A W L V K G G G P S S G A P P P S C 　
44 H X Q G T F T S D V S S Y L L G Q Q Q K E F I A W L V K G CH X Q G T F T S D V S S Y L L G Q Q Q K E F I A W L V K G C 　
55 H X Q G T F T S D V S S Y L L G Q Q Q K E F I A W L V K G G G P S S G A P P P S CH X Q G T F T S D V S S Y L L G Q Q Q K E F I A W L V K G G G P S S G A P P P S C 　
66 H X Q G T F T S D V S S Y L D G Q A A K E F V A W L L K G CH X Q G T F T S D V S S Y L D G Q A A K E F V A W L L K G C 　
77 H X Q G T F T S D V S K Y L D G Q A A K E F V A W L L K G C H X Q G T F T S D V S K Y L D G Q A A K E F V A W L L K G C 　
88 H X Q G T F T S D V S K Y L D G Q A A Q E F V A W L L K G CH X Q G T F T S D V S K Y L D G Q A A Q E F V A W L L K G C 　
99 H X Q G T F T S D V S K Y L D G Q A A Q E F V A W L L A G C H X Q G T F T S D V S K Y L D G Q A A Q E F V A W L L A G C 　
1010 H X Q G T F T S D V S K Y L D G Q A A Q E F V A W L L A G G G P S S G A P P P S CH X Q G T F T S D V S K Y L D G Q A A Q E F V A W L L A G G G P S S G A P P P S C 　
1111 CA G E G T F T S D L S K Y L D S R R Q Q L F V Q W L K A G G P S S G A P P P S H G CA G E G T F T S D L S K Y L D S R R Q Q L F V Q W L K A G G P S S G A P P P S H G 　
1212 CA G E G T F I S D L S K Y M D E Q A V Q L F V E W L M A G G P S S G A P P P S H GCA G E G T F I S D L S K Y M D E Q A V Q L F V E W L M A G G P S S G A P P P S H G 　
1313 CA G E G T F I S D Y S I Q L D E I A V Q D F V E W L L A Q K P S S G A P P P S H GCA G E G T F I S D Y S I Q L D E I A V Q D F V E W L L A Q K P S S G A P P P S H G 　
1414 CA G Q G T F T S D Y S I Q L D E I A V R D F V E W L K N G G P S S G A P P P S H G CA G Q G T F T S D Y S I Q L D E I A V R D F V E W L K N G G P S S G A P P P S H G 　
1515 CA G Q G T F T S D L S K Q M D E E A V R L F I E W L K N G G P S S G A P P P S H GCA G Q G T F T S D L S K Q M D E E A V R L F I E W L K N G G P S S G A P P P S H G 　
1616 CA G Q G T F T S D L S K Q M D S E A Q Q L F I E W L K N G G P S S G A P P P S H GCA G Q G T F T S D L S K Q M D S E A Q Q L F I E W L K N G G P S S G A P P P S H G 　
1717 CA G Q G T F T S D L S K Q M D E E R A R E F I E W L L A Q K P S S G A P P P S H GCA G Q G T F T S D L S K Q M D E E R A R E F I E W L L A Q K P S S G A P P P S H G 　
1818 CA G Q G T F T S D L S K Q M D S E R A R E F I E W L K N T G P S S G A P P P S H G CA G Q G T F T S D L S K Q M D S E R A R E F I E W L K N T G P S S G A P P P S H G 　
1919 CA G Q G T F T S D L S I Q Y D S E H Q R D F I E W L K D T G P S S G A P P P S H GCA G Q G T F T S D L S I Q Y D S E H Q R D F I E W L K D T G P S S G A P P P S H G 　
2020 CA G Q G T F T S D L S I Q Y E E E A Q Q D F V E W L K D T G P S S G A P P P S H GCA G Q G T F T S D L S I Q Y E E E A Q Q D F V E W L K D T G P S S G A P P P S H G 　
2121 Y X Q G T F T S D Y S K Y L D E C R A K E F V Q W L L D H H P S S G Q P P P S YXQGTFTSDYSKYLD E CRA K EFVQWLLDHHPSSGQPPPS 고리 형성ring formation
2222 Y X Q G T F T S D Y S K C L D E K R A K E F V Q W L L D H H P S S G Q P P P SYXQGTFTSDYSKCLD E KRA K EFVQWLLDHHPSSGQPPPS 고리 형성ring formation
2323 Y X Q G T F T S D Y S K Y L D E C R A K E F V Q W L L A Q K G K K N D W K H N I T YXQGTFTSDYSKYLD E CRA K EFVQWLLAQKGKKKNDWKHNIT 고리 형성ring formation
2424 Y X Q G T F T S D Y S K Y L D E C R A K E F V Q W L K N G G P S S G A P P P SYXQGTFTSDYSKYLD E CRA K EFVQWLKNGGPSSGAPPPS 고리 형성ring formation
2525 H X Q G T F T S D C S K Y L D E R A A Q D F V Q W L L D G G P S S G A P P P S H X Q G T F T S D C S K Y L D E R A A Q D F V Q W L L D G G P S S G A P P P S 　
2626 H X Q G T F T S D C S K Y L D S R A A Q D F V Q W L L D G G P S S G A P P P S H X Q G T F T S D C S K Y L D S R A A Q D F V Q W L L D G G P S S G A P P P S 　
2727 H X Q G T F T S D Y S K Y L D E R A C Q D F V Q W L L D Q G G P S S G A P P P S H X Q G T F T S D Y S K Y L D E R A C Q D F V Q W L L D Q G G P S S G A P P P S 　
2828 H X Q G T F T S D Y S K Y L D E K R A Q E F V C W L L A Q K G K K N D W K H N I T H X Q G T F T S D Y S K Y L D E K R A Q E F V C W L L A Q K G K K N D W K H N I T 　
2929 H X Q G T F T S D Y S K Y L D E K A A K E F V Q W L L N T C HXQGTFTSDYSKYLD E KAA K EFVQWLLNTC 고리 형성ring formation
3030 H X Q G T F T S D Y S K Y L D E K A Q K E F V Q W L L D T CHXQGTFTSDYSKYLD E KAQ K EFVQWLLDTC 고리 형성ring formation
3131 H X Q G T F T S D Y S K Y L D E K A C K E F V Q W L L A Q HXQGTFTSDYSKYLD E KAC K EFVQWLLAQ 고리 형성ring formation
3232 H X Q G T F T S D Y S K Y L D E K A C K D F V Q W L L D G G P S S G A P P P S HXQGTFTSDYSKYLD E KAC K DFVQWLLDGGPSSGAPPPS 고리 형성ring formation
3333 H X Q G T F T S D Y S I A M D E I H Q K D F V N W L L A Q K C HXQGTFTSDYSIAMD E IHQ K DFVNWLLAQKC 고리 형성ring formation
3434 H X Q G T F T S D Y S K Y L D E K R Q K E F V N W L L A Q K C HXQGTFTSDYSKYLD E KRQ K EFVNWLLAQKC 고리 형성ring formation
3535 H X Q G T F T S D Y S I A M D E I H Q K D F V N W L L N T K C HXQGTFTSDYSIAMD E IHQ K DFVNWLLNTKC 고리 형성ring formation
3636 H X Q G T F T S D Y S K Y L C E K R Q K E F V Q W L L N G G P S S G A P P P S G HXQGTFTSDYSKYLC E KRQ K EFVQWLLNGGPSSGAPPPSG 고리 형성ring formation
3737 H X Q G T F T S D Y S K Y L D E C R Q K E F V Q W L L N G G P S S G A P P P S G HXQGTFTSDYSKYLD E CRQ K EFVQWLLNGGPSSGAPPPSG 고리 형성ring formation
3838 CA X Q G T F T S D K S S Y L D E R A A Q D F V Q W L L D G G P S S G A P P P S SCA X Q G T F T S D K S S Y L D E R A A Q D F V Q W L L D G G P S S G A P P P S S 　
3939 H X Q G T F T S D Y S K Y L D G Q H A Q C F V A W L L A G G G P S S G A P P P S H X Q G T F T S D Y S K Y L D G Q H A Q C F V A W L L A G G G P S S G A P P P S 　
4040 H X Q G T F T S D K S K Y L D E R A C Q D F V Q W L L D G G P S S G A P P P S H X Q G T F T S D K S K Y L D E R A C Q D F V Q W L L D G G P S S G A P P P S 　
4141 H X Q G T F T S D K S K Y L D E C A A Q D F V Q W L L D G G P S S G A P P P SH X Q G T F T S D K S K Y L D E C A A Q D F V Q W L L D G G P S S G A P P P S 　
4242 Y X Q G T F T S D Y S K Y L D E K R A K E F V Q W L L D H H P S S G Q P P P S C YXQGTFTSDYSKYLD E KRA K EFVQWLLDHHPSSGQPPPSC 고리 형성ring formation
4343 Y X Q G T F T S D Y S K Y L D E K R A K E F V Q W L L D H H C S S G Q P P P S YXQGTFTSDYSKYLD E KRA K EFVQWLLDHHCSSGQPPPS 고리 형성ring formation
4444 H G Q G T F T S D C S K Q L D G Q A A Q E F V A W L L A G G P S S G A P P P S H G Q G T F T S D C S K Q L D G Q A A Q E F V A W L L A G G P S S G A P P P S 　
4545 H G Q G T F T S D C S K Y M D G Q A A Q D F V A W L L A G G P S S G A P P P S H G Q G T F T S D C S K Y M D G Q A A Q D F V A W L L A G G P S S G A P P P S 　
4646 H G Q G T F T S D C S K Y L D E Q H A Q E F V A W L L A G G P S S G A P P P S H G Q G T F T S D C S K Y L D E Q H A Q E F V A W L L A G G P S S G A P P P S 　
4747 H G Q G T F T S D C S K Y L D G Q R A Q E F V A W L L A G G P S S G A P P P S H G Q G T F T S D C S K Y L D G Q R A Q E F V A W L L A G G P S S G A P P P S 　
4848 H G Q G T F T S D C S K Y L D G Q R A Q D F V N W L L A G G P S S G A P P P S H G Q G T F T S D C S K Y L D G Q R A Q D F V N W L L A G G P S S G A P P P S 　
4949 CA X Q G T F T S D Y S I C M D E I H Q K D F V N W L L N T K CA XQGTFTSDYSICMD E IHQ K DFVNWLLNTK 고리 형성ring formation
5050 H X Q G T F T S D Y S K Y L D E K R A K E F V Q W L L D H H P S S G Q P P P S C HXQGTFTSDYSKYLD E KRA K EFVQWLLDHHPSSGQPPPSC 고리 형성ring formation
5151 H X Q G T F T S D Y S K Y L D E K R Q K E F V Q W L L N T C HXQGTFTSDYSKYLD E KRQ K EFVQWLLNTC 고리 형성ring formation
5252 H X Q G T F T S D Y S K Y L D E K R Q K E F V Q W L L D T C HXQGTFTSDYSKYLD E KRQ K EFVQWLLDTC 고리 형성ring formation
5353 H X E G T F T S D Y S I A M D E I H Q K D F V N W L L A Q CHXEGTFTSDYSIAMD E IHQ K DFVNWLLAQC 고리 형성ring formation
5454 H X E G T F T S D Y S I A M D E I H Q K D F V D W L L A E C HXEGTFTSDYSIAMD E IHQ K DFVDWLLAEC 고리 형성ring formation
5555 H X Q G T F T S D Y S I A M D E I H Q K D F V N W L L A Q C HXQGTFTSDYSIAMD E IHQ K DFVNWLLAQC 고리 형성ring formation
5656 H X Q G T F T S D Y S K Y L D E K R Q K E F V N W L L A Q C HXQGTFTSDYSKYLD E KRQ K EFVNWLLAQC 고리 형성ring formation
5757 H X Q G T F T S D Y S I A M D E I H Q K D F V N W L L N T C HXQGTFTSDYSIAMD E IHQ K DFVNWLLNTC 고리 형성ring formation
5858 H X Q G T F T S D Y S K Y L D E K R Q K E F V Q W L L N T K C HXQGTFTSDYSKYLD E KRQ K EFVQWLLNTKC 고리 형성ring formation
5959 CA X Q G T F T S D Y S I C M D E K H Q K D F V N W L L N T K CA XQGTFTSDYSICMD E KHQ K DFVNWLLNTK 고리 형성ring formation
6060 CA X Q G T F T S D Y S I A M D E K H C K D F V N W L L N T K CA XQGTFTSDYSIAMD E KHC K DFVNWLLNTK 고리 형성ring formation
6161 CA X Q G T F T S D Y S I A M D E I A C K D F V N W L L N T K CA XQGTFTSDYSIAMD E IAC K DFVNWLLNTK 고리 형성ring formation
6262 CA X Q G T F T S D K S K Y L D E R A A Q D F V Q W L L D G G P S S G A P P P SCA X Q G T F T S D K S K Y L D E R A A Q D F V Q W L L D G G P S S G A P P P S
6363 CA X Q G T F T S D C S K Y L D E R A A Q D F V Q W L L D G G P S S G A P P P SCA X Q G T F T S D C S K Y L D E R A A Q D F V Q W L L D G G P S S G A P P P S
6464 Y X Q G T F T S D Y S K Y L D E C A A K E F V Q W L L D H H P S S G Q P P P SYXQGTFTSDYSKYLD E CAA K EFVQWLLDHHPSSGQPPPS 고리 형성　ring formation
6565 H X Q G T F T S D Y S K C L D E K R A K E F V Q W L L D H H P S S G Q P P P S HXQGTFTSDYSKCLD E KRA K EFVQWLLDHHPSSGQPPPS 고리 형성ring formation
6666 Y X Q G T F T S D Y S K Y L D E C R A K D F V Q W L L D H H P S S G Q P P P S YXQGTFTSDYSKYLD E CRA K DFVQWLLDHHPSSGQPPPS 고리 형성ring formation
6767 Y X Q G T F T S D Y S K Y L D E C A A K D F V Q W L L D H H P S S G Q P P P S YXQGTFTSDYSKYLD E CAA K DFVQWLLDHHPSSGQPPPS 고리 형성ring formation
6868 Y X Q G T F T S D Y S K C L D E K A A K E F V Q W L L D H H P S S G Q P P P SYXQGTFTSDYSKCLD E KAA K EFVQWLLDHHPSSGQPPPS 고리 형성ring formation
6969 Y X Q G T F T S D Y S K C L D E R A A K E F V Q W L L D H H P S S G Q P P P S YXQGTFTSDYSKCLD E RAA K EFVQWLLDHHPSSGQPPPS 고리 형성ring formation
7070 Y X Q G T F T S D Y S K C L D E K R A K D F V Q W L L D H H P S S G Q P P P S YXQGTFTSDYSKCLD E KRA K DFVQWLLDHHPSSGQPPPS 고리 형성ring formation
7171 Y X Q G T F T S D Y S K Y L D E R A C K D F V Q W L L D H H P S S G Q P P P S YXQGTFTSDYSKYLD E RAC K DFVQWLLDHHPSSGQPPPS 고리 형성ring formation
7272 Y X Q G T F T S D C S K Y L D E R A A K D F V Q W L L D H H P S S G Q P P P S YXQGTFTSDCSKYLD E RAA K DFVQWLLDHHPSSGQPPPS 고리 형성ring formation
7373 CA X Q G T F T S D Y S K Y L D E C R A K E F V Q W L L D H H P S S G Q P P P S CA XQGTFTSDYSKYLD E CRA K EFVQWLLDHHPSSGQPPPS 고리 형성ring formation
7474 CA X Q G T F T S D Y S K C L D E K R A K E F V Q W L L D H H P S S G Q P P P S CA XQGTFTSDYSKCLD E KRA K EFVQWLLDHHPSSGQPPPS 고리 형성ring formation
7575 Y X Q G T F T S D Y S K Y L D E K A A K E F V Q W L L D H H P S S G Q P P P S C YXQGTFTSDYSKYLD E KAA K EFVQWLLDHHPSSGQPPPSC 고리 형성ring formation
7676 Y X Q G T F T S D Y S K Y L D E K R A K D F V Q W L L D H H P S S G Q P P P S CYXQGTFTSDYSKYLD E KRA K DFVQWLLDHHPSSGQPPPSC 고리 형성ring formation
7777 Y X Q G T F T S D Y S K Y L D E K A A K D F V Q W L L D H H P S S G Q P P P S C YXQGTFTSDYSKYLD E KAA K DFVQWLLDHHPSSGQPPPSC 고리 형성ring formation
7878 H X Q G T F T S D Y S K Y L D E K R Q K E F V Q W L L D T K C HXQGTFTSDYSKYLD E KRQ K EFVQWLLDTKC 고리 형성ring formation
7979 H X E G T F T S D Y S I A M D E I H Q K D F V N W L L A Q K C HXEGTFTSDYSIAMD E IHQ K DFVNWLLAQKC 고리 형성ring formation
8080 H X E G T F T S D Y S I A M D E I H Q K D F V D W L L A E K C HXEGTFTSDYSIAMD E IHQ K DFVDWLLAEKC 고리 형성ring formation
8181 CA X Q G T F T S D Y S K Y L D E K R Q K E F V Q W L L N T CCA XQGTFTSDYSKYLD E KRQ K EFVQWLLNTC 고리 형성ring formation
8282 CA X Q G T F T S D Y S K Y L D E K R Q K E F V Q W L L D T C CA XQGTFTSDYSKYLD E KRQ K EFVQWLLDTC 고리 형성ring formation
8383 CA X E G T F T S D Y S I A M D E I H Q K D F V N W L L A Q C CA XEGTFTSDYSIAMD E IHQ K DFVNWLLAQC 고리 형성ring formation
8484 CA X E G T F T S D Y S I A M D E I H Q K D F V D W L L A E C CA XEGTFTSDYSIAMD E IHQ K DFVDWLLAEC 고리 형성ring formation
8585 CA X Q G T F T S D Y S I A M D E I H Q K D F V N W L L A Q C CA XQGTFTSDYSIAMD E IHQ K DFVNWLLAQC 고리 형성ring formation
8686 CA X Q G T F T S D Y S K Y L D E K R Q K E F V N W L L A Q C CA XQGTFTSDYSKYLD E KRQ K EFVNWLLAQC 고리 형성ring formation
8787 CA X Q G T F T S D Y S I A M D E I H Q K D F V N W L L N T CCA XQGTFTSDYSIAMD E IHQ K DFVNWLLNTC 고리 형성ring formation
8888 CA X Q G T F T S D Y S K Y L D E K R Q K E F V Q W L L N T K C CA XQGTFTSDYSKYLD E KRQ K EFVQWLLNTKC 고리 형성ring formation
8989 CA X Q G T F T S D Y S K Y L D E K R Q K E F V Q W L L D T K C CA XQGTFTSDYSKYLD E KRQ K EFVQWLLDTKC 고리 형성ring formation
9090 CA X E G T F T S D Y S I A M D E I H Q K D F V N W L L A Q K CCA XEGTFTSDYSIAMD E IHQ K DFVNWLLAQKC 고리 형성ring formation
9191 CA X E G T F T S D Y S I A M D E I H Q K D F V D W L L A E K CCA XEGTFTSDYSIAMD E IHQ K DFVDWLLAEKC 고리 형성ring formation
9292 CA X Q G T F T S D Y S I A M D E I H Q K D F V N W L L A Q K C CA XQGTFTSDYSIAMD E IHQ K DFVNWLLAQKC 고리 형성ring formation
9393 CA X Q G T F T S D Y S K Y L D E K R Q K E F V N W L L A Q K C CA XQGTFTSDYSKYLD E KRQ K EFVNWLLAQKC 고리 형성ring formation
9494 CA X Q G T F T S D Y S I A M D E I H Q K D F V N W L L N T K CCA XQGTFTSDYSIAMD E IHQ K DFVNWLLNTKC 고리 형성ring formation
9595 Y X Q G T F T S D Y S K Y L D E K R A K E F V Q W L L C H H P S S G Q P P P S YXQGTFTSDYSKYLD E KRA K EFVQWLLCHHPSSGQPPPS 고리 형성ring formation
9696 Y X Q G T F T S D Y S K Y L D E K R A K E F V Q W L L D H C P S S G Q P P P SYXQGTFTSDYSKYLD E KRA K EFVQWLLDHCPSSGQPPPS 고리 형성ring formation
9797 Y X Q G T F T S D Y S K Y L D E K R A K E F V Q W L L D C H P S S G Q P P P SYXQGTFTSDYSKYLD E KRA K EFVQWLLDCHPSSGQPPPS 고리 형성ring formation
9898 Y X Q G T F T S D Y S K A L D E K A A K E F V N W L L D H H P S S G Q P P P S CYXQGTFTSDYSKALD E KAA K EFVNWLLDHHPSSGQPPPSC 고리 형성ring formation
9999 Y X Q G T F T S D Y S K A L D E K A A K D F V N W L L D H H P S S G Q P P P S CYXQGTFTSDYSKALD E KAA K DFVNWLLDHHPSSGQPPPSC 고리 형성ring formation
100100 Y X Q G T F T S D Y S K A L D E K A A K E F V Q W L L D Q H P S S G Q P P P S CYXQGTFTSDYSKALD E KAA K EFVQWLLDQHPSSGQPPPSC 고리 형성ring formation
101101 Y X Q G T F T S D Y S K A L D E K A A K E F V N W L L D Q H P S S G Q P P P S C YXQGTFTSDYSKALD E KAA K EFVNWLLDQHPSSGQPPPSC 고리 형성ring formation
102102 Y X Q G T F T S D Y S K A L D E K A A K D F V N W L L D Q H P S S G Q P P P S C YXQGTFTSDYSKALD E KAA K DFVNWLLDQHPSSGQPPPSC 고리 형성ring formation
상기 표 1에 기재된 서열에서 X로 표기된 아미노산은 비천연형 아미노산인 Aib (2-아미노이소부티르산(2-aminoisobutyric acid))이며, 아미노산 기호 아래 밑줄은 밑줄 친 해당 아미노산 쌍의 측쇄들 사이에 락탐 고리가 형성된 것을 가리킨다. 또한, 상기 표 1에서 CA는 4-이미다조아세틸(4-imidazoacetyl)을 의미한다.The amino acid denoted by X in the sequence shown in Table 1 is Aib (2-aminoisobutyric acid), a non-natural amino acid, and the underlined under the amino acid symbol is the lactam ring between the side chains of the underlined amino acid pair. indicates that it is formed. In addition, in Table 1, CA means 4-imidazoacetyl (4-imidazoacetyl).
실시예 2: 삼중 활성체의 지속형 결합체 제조Example 2: Preparation of long-acting conjugates of triple activators
양 말단에 각각 말레이미드기 및 알데히드기를 가지는 10kDa의 PEG, 즉 말레이미드-PEG-알데히드 (10kDa, NOF, 일본)를 실시예 1의 삼중활성체 (서열번호 21, 22, 42, 43, 50, 77, 및 96)의 시스테인 잔기에 페길화시키기 위하여, 삼중활성체와 말레이미드-PEG-알데히드의 몰비를 1 : 1 내지 3, 단백질의 농도를 1 내지 5 ㎎/㎖로 하여 저온에서 0.5 내지 3 시간 동안 반응시켰다. 이때, 반응은 50 mM Tris 완충액(pH 7.5)에 20 내지 60% 아이소프로판올이 첨가된 환경 하에서 수행되었다. 구체적으로, 삼중활성체와 폴리에틸렌글리콜 링커의 연결체를 제조하기 위하여 양 말단의 수소가 각각 3-(3-말레이미도프로피온아미도)프로필기 및 3-옥소프로필기(프로피온알데히드기)로 치환된 분자량 10 kDa의 선형 개질 폴리에틸렌글리콜인 말레이미드-PEG-알데히드(일본 NOF사)를 상기 삼중활성체의 시스테인 잔기에 반응시킴으로써 삼중활성체를 말레이미드-PEG-알데히드의 말레이미드쪽 말단에 페길화시켰다.The triple activator of Example 1 (SEQ ID NOs: 21, 22, 42, 43, 50, 77, and 96) for pegylation to the cysteine residues, the molar ratio of the triple activator to maleimide-PEG-aldehyde is 1:1 to 3, and the protein concentration is 1 to 5 mg/ml, and 0.5 to 3 at low temperature. reacted for an hour. At this time, the reaction was carried out in an environment in which 20 to 60% isopropanol was added to 50 mM Tris buffer (pH 7.5). Specifically, the molecular weight in which the hydrogens at both terminals are substituted with 3-(3-maleimidopropionamido)propyl groups and 3-oxopropyl groups (propionaldehyde groups), respectively, to prepare a linker between the triple activator and the polyethylene glycol linker. The triple activator was pegylated to the maleimide end of maleimide-PEG-aldehyde by reacting a 10 kDa linearly modified polyethylene glycol, maleimide-PEG-aldehyde (NOF, Japan) with the cysteine residue of the triple activator.
반응이 종료된 후, 상기 반응액을 SP 세파로스 HP (GE healthcare, 미국)에 적용하여 시스테인에 모노-페길화된 삼중활성체를 정제하였다.After the reaction was completed, the reaction solution was applied to SP Sepharose HP (GE healthcare, USA) to purify the tri-activator mono-pegylated to cysteine.
다음으로, 상기 정제된 모노-페길화된 삼중활성체와 면역글로불린 Fc (서열번호 138의 동종이량체)를 몰비를 1 : 1 내지 5, 단백질의 농도를 10 내지 50㎎/㎖로 하여 4 내지 8℃에서 12 내지 18시간 동안 반응시켰다. 반응은 100 mM 인산칼륨 완충액 (pH 6.0)에 환원제인 10 내지 50 mM 소디움시아노보로하이드라이드와 10 내지 30 % 아이소프로판올이 첨가된 환경 하에서 수행되었다. 반응이 종료된 후, 상기 반응액을 부틸 세파로스 FF 정제컬럼 (GE healthcare, 미국)과 Source ISO 정제컬럼 (GE healthcare, 미국)에 적용하여, 삼중활성체와 면역글로불린 Fc를 포함하는 결합체를 정제하였다. 정제된 이 지속형 결합체는 분자 내에서 삼중활성체 펩타이드, 폴리에틸렌 글리콜(PEG) 링커와 Fc 이량체가 1:1:1의 몰 비로 공유결합 연결된 구조이며, PEG 링커는 Fc 이량체의 두 폴리펩타이드 사슬 중 한 사슬에만 연결되어 있다.Next, the purified mono-pegylated tri-activator and immunoglobulin Fc (homodimer of SEQ ID NO: 138) were mixed in a molar ratio of 1:1 to 5 and a protein concentration of 10 to 50 mg/ml from 4 to The reaction was carried out at 8°C for 12 to 18 hours. The reaction was carried out in an environment in which 10 to 50 mM sodium cyanoborohydride and 10 to 30% isopropanol as reducing agents were added to 100 mM potassium phosphate buffer (pH 6.0). After completion of the reaction, the reaction solution was applied to the butyl sepharose FF purification column (GE healthcare, USA) and the Source ISO purification column (GE healthcare, USA) to purify the conjugate containing the triple activator and immunoglobulin Fc. did This purified long-acting conjugate has a structure in which a triple active peptide, a polyethylene glycol (PEG) linker and an Fc dimer are covalently linked in a molar ratio of 1:1:1, and the PEG linker is two polypeptide chains of an Fc dimer. It is connected to only one of the chains.
한편, 상기 면역글로불린 Fc는 서열번호 138의 아미노산 서열 (221개의 아미노산으로 구성됨)을 갖는 단량체 2개가 각 단량체의 3번 아미노산인 시스테인 사이에 이황화 결합을 통해 동종이량체를 형성하고, 상기 동종이량체의 단량체는 각각 독립적으로 35번 및 95번의 시스테인 간의 내부의 이황화 결합 및 141번 및 199번의 시스테인 간의 내부의 이황화 결합이 형성된 것이다. 구체적으로 면역글로불린 Fc는 N-말단에 Pro-Ser-Cys-Pro 서열의 힌지 영역을 갖는 면역글로불린 Fc 절편(서열번호 138의 사슬 두 개가 이황화결합으로 연결된 동종이량체)을 사용하여 국제공개특허 WO2007/021129에 기재된 방법으로 제조하였다.On the other hand, in the immunoglobulin Fc, two monomers having the amino acid sequence of SEQ ID NO: 138 (consisting of 221 amino acids) form a homodimer through a disulfide bond between cysteine, which is the 3rd amino acid of each monomer, and the homodimer The monomers of are each independently formed with an internal disulfide bond between cysteines at positions 35 and 95 and an internal disulfide bond between cysteines at positions 141 and 199. Specifically, the immunoglobulin Fc is an immunoglobulin Fc fragment (a homodimer in which two chains of SEQ ID NO: 138 are linked by a disulfide bond) having a hinge region of the Pro-Ser-Cys-Pro sequence at the N-terminus. International Patent Publication No. WO2007 /021129 was prepared by the method described.
제조 후 역상 크로마토그래피, 크기배제 크로마토그래피 및 이온교환 크로마토그래피로 분석한 순도는 95 % 이상이었다.After preparation, the purity analyzed by reverse phase chromatography, size exclusion chromatography, and ion exchange chromatography was 95% or more.
여기서, 서열번호 21의 삼중활성체 및 면역글로불린 Fc가 PEG를 통하여 연결된 결합체를, '서열번호 21과 면역글로불린 Fc를 포함하는 결합체' 혹은 '서열번호 21의 지속형 결합체'로 명명하였고, 이들은 본원에서 혼용되어 사용될 수 있다. Here, the conjugate in which the triple activator of SEQ ID NO: 21 and the immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 21 and immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 21'. may be used interchangeably.
여기서, 서열번호 22의 삼중활성체 및 면역글로불린 Fc가 PEG를 통하여 연결된 결합체를, '서열번호 22와 면역글로불린 Fc를 포함하는 결합체' 혹은 '서열번호 22의 지속형 결합체'로 명명하였고, 이들은 본원에서 혼용되어 사용될 수 있다.Herein, the conjugate in which the triple activator of SEQ ID NO: 22 and immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 22 and immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 22'. may be used interchangeably.
여기서, 서열번호 42의 삼중활성체 및 면역글로불린 Fc가 PEG를 통하여 연결된 결합체를, '서열번호 42와 면역글로불린 Fc를 포함하는 결합체' 혹은 '서열번호 42의 지속형 결합체'로 명명하였고, 이들은 본원에서 혼용되어 사용될 수 있다. Here, the conjugate in which the triple activator of SEQ ID NO: 42 and the immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 42 and an immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 42'. may be used interchangeably.
여기서, 서열번호 43의 삼중활성체 및 면역글로불린 Fc가 PEG를 통하여 연결된 결합체를, '서열번호 43과 면역글로불린 Fc를 포함하는 결합체' 혹은 '서열번호 43의 지속형 결합체'로 명명하였고, 이들은 본원에서 혼용되어 사용될 수 있다.Here, the conjugate in which the triple activator of SEQ ID NO: 43 and the immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 43 and an immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 43', which may be used interchangeably.
여기서, 서열번호 50의 삼중활성체 및 면역글로불린 Fc가 PEG를 통하여 연결된 결합체를, '서열번호 50과 면역글로불린 Fc를 포함하는 결합체' 혹은 '서열번호 50의 지속형 결합체'로 명명하였고, 이들은 본원에서 혼용되어 사용될 수 있다. Here, the conjugate in which the triple activator of SEQ ID NO: 50 and immunoglobulin Fc are linked via PEG was named as 'a conjugate comprising SEQ ID NO: 50 and immunoglobulin Fc' or 'a long-acting conjugate of SEQ ID NO: 50', may be used interchangeably.
여기서, 서열번호 77의 삼중활성체 및 면역글로불린 Fc가 PEG를 통하여 연결된 결합체를, '서열번호 77와 면역글로불린 Fc를 포함하는 결합체' 혹은 '서열번호 77의 지속형 결합체'로 명명하였고, 이들은 본원에서 혼용되어 사용될 수 있다.Here, the conjugate in which the triple activator of SEQ ID NO: 77 and the immunoglobulin Fc are linked via PEG was designated as a 'conjugate comprising SEQ ID NO: 77 and immunoglobulin Fc' or a 'persistent conjugate of SEQ ID NO: 77' may be used interchangeably.
여기서, 서열번호 96의 삼중활성체 및 면역글로불린 Fc가 PEG를 통하여 연결된 결합체를, '서열번호 96과 면역글로불린 Fc를 포함하는 결합체' 혹은 '서열번호 96의 지속형 결합체'로 명명하였고, 이들은 본원에서 혼용되어 사용될 수 있다.Here, the conjugate in which the triple activator of SEQ ID NO: 96 and the immunoglobulin Fc are linked through PEG was designated as a 'conjugate comprising SEQ ID NO: 96 and immunoglobulin Fc' or a 'continuous conjugate of SEQ ID NO: 96'. may be used interchangeably.
실시예 3: 삼중 활성체 및 이의 지속형 결합체의Example 3: Triple activator and long-acting conjugate thereof in vitro in vitro 활성 측정 activity measurement
상기 실시예 1 및 2에서 제조된 삼중활성체와 이의 지속형 결합체의 활성을 측정하기 위해 GLP-1 수용체, 글루카곤(GCG) 수용체, 및 GIP 수용체가 각각 형질전환된 세포주를 이용하여 in vitro에서 세포 활성을 측정하는 방법을 이용하였다.In order to measure the activity of the triple activator prepared in Examples 1 and 2 and the long-acting conjugate thereof, cells were in vitro using cell lines transformed with GLP-1 receptor, glucagon (GCG) receptor, and GIP receptor, respectively. A method for measuring activity was used.
상기 각 세포주는 CHO (chinese hamster ovary)에 인간 GLP-1 수용체, 인간 GCG 수용체 및 인간 GIP 수용체 유전자를 각각 발현하도록 형질 전환된 것으로서, GLP-1, GCG 및 GIP의 활성을 측정하기에 적합하다. 따라서, 각 부분에 대한 활성을 각각의 형질 전환 세포주를 이용하여 측정하였다.Each of the above cell lines was transformed to express human GLP-1 receptor, human GCG receptor and human GIP receptor genes in Chinese hamster ovary (CHO), respectively, and is suitable for measuring the activities of GLP-1, GCG and GIP. Therefore, the activity for each part was measured using each transformed cell line.
상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체의 GLP-1 활성 측정을 위해 인간 GLP-1을 50nM 부터 4배씩 0.000048nM까지 연속적으로 희석하고, 상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체를 400nM 부터 4배씩 0.00038nM까지 연속적으로 희석하였다. 상기 배양된 인간 GLP-1 수용체가 발현된 CHO 세포에서 배양액을 제거하고 연속적으로 희석된 각 물질들을 5㎕씩 상기 세포에 첨가한 다음, cAMP 항체가 포함된 완충액을 5㎕씩 추가 한 뒤 15분 동안 상온에서 배양하였다. 그런 다음 세포용해완충액 (cell lysis buffer)이 포함된 detection mix를 10㎕씩 가하여 세포를 용해시키고, 90분 동안 상온에서 반응시켰다. 상기 반응이 완료된 세포용해물을 LANCE cAMP kit (PerkinElmer, USA)에 적용하여 축적된 cAMP를 통해 EC50값을 산출한 후, 상호 비교하였다. 인간 GLP-1 대비 상대 역가는 하기 표 2와 표 3에 나타내었다.Human GLP-1 was serially diluted from 50 nM to 0.000048 nM by 4 times to measure the GLP-1 activity of the triple activator and its long-acting conjugate prepared in Examples 1 and 2, and prepared in Examples 1 and 2 The triple activator and its long-acting conjugate were serially diluted from 400 nM to 0.00038 nM by 4 times. The culture medium was removed from the cultured CHO cells expressing the human GLP-1 receptor, 5 μl of each serially diluted substance was added to the cells, and then 5 μl of a buffer containing cAMP antibody was added for 15 minutes. during incubation at room temperature. Then, 10 μl of a detection mix containing cell lysis buffer was added to lyse the cells, followed by reaction at room temperature for 90 minutes. The cell lysate after the reaction was completed was applied to the LANCE cAMP kit (PerkinElmer, USA) to calculate the EC 50 value through the accumulated cAMP, and then compared with each other. Relative titers compared to human GLP-1 are shown in Tables 2 and 3 below.
상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체의 GCG 활성 측정을 위해 인간 GCG을 50nM 부터 4배씩 0.000048nM까지 연속적으로 희석하고, 상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체를 400nM 부터 4배씩 0.00038nM까지 연속적으로 희석하였다. 상기 배양된 인간 GCG 수용체가 발현된 CHO 세포에서 배양액을 제거하고 연속적으로 희석된 각 물질들을 5㎕씩 상기 세포에 첨가한 다음, cAMP 항체가 포함된 완충액을 5㎕씩 추가 한 뒤 15분 동안 상온에서 배양하였다. 그런 다음 세포용해완충액(cell lysis buffer)이 포함된 detection mix를 10㎕씩 가하여 세포를 용해시키고, 90분 동안 상온에서 반응시켰다. 상기 반응이 완료된 세포용해물을 LANCE cAMP kit (PerkinElmer, USA)에 적용하여 축적된 cAMP를 통해 EC50값을 산출한 후, 상호 비교하였다. 인간 GCG 대비 상대 역가는 하기 표2와 표3에 나타내었다.In order to measure the GCG activity of the triple activator prepared in Examples 1 and 2 and the long-acting conjugate thereof, human GCG was serially diluted from 50 nM to 0.000048 nM by 4 times, and the triple activator prepared in Examples 1 and 2 was serially diluted. and its long-acting conjugate were serially diluted from 400 nM to 0.00038 nM in 4-fold increments. The culture medium was removed from the cultured CHO cells expressing human GCG receptor, 5 μl of each serially diluted substance was added to the cells, and 5 μl of a buffer containing cAMP antibody was added thereto, followed by room temperature for 15 minutes. cultured in Then, 10 μl of a detection mix containing a cell lysis buffer was added to lyse the cells, followed by reaction at room temperature for 90 minutes. The cell lysate after the reaction was completed was applied to the LANCE cAMP kit (PerkinElmer, USA) to calculate the EC 50 value through the accumulated cAMP, and then compared with each other. Relative titers compared to human GCG are shown in Tables 2 and 3 below.
상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체의 GIP 활성 측정을 위해 인간 GIP을 50nM 부터 4배씩 0.000048nM까지 연속적으로 희석하고, 상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체를 400nM 부터 4배씩 0.00038nM까지 연속적으로 희석하였다. 상기 배양된 인간 GIP 수용체가 발현된 CHO 세포에서 배양액을 제거하고 연속적으로 희석된 각 물질들을 5㎕씩 상기 세포에 첨가한 다음, cAMP 항체가 포함된 완충액을 5㎕씩 추가 한 뒤 15분 동안 상온에서 배양하였다. 그런 다음 세포용해완충액(cell lysis buffer)이 포함된 detection mix를 10㎕씩 가하여 세포를 용해시키고, 90분 동안 상온에서 반응시켰다. 상기 반응이 완료된 세포용해물을 LANCE cAMP kit (PerkinElmer, USA)에 적용하여 축적된 cAMP를 통해 EC50값을 산출한 후, 상호 비교하였다. 인간 GIP 대비 상대 역가는 하기 표 2와 표 3에 나타내었다.In order to measure the GIP activity of the triple activator prepared in Examples 1 and 2 and its long-acting conjugate, human GIP was serially diluted from 50 nM to 0.000048 nM by 4 times, and the triple activator prepared in Examples 1 and 2 was serially diluted. and its long-acting conjugate were serially diluted from 400 nM to 0.00038 nM in 4-fold increments. The culture medium was removed from the cultured CHO cells expressing the human GIP receptor, 5 μl of each serially diluted substance was added to the cells, and 5 μl of a buffer containing cAMP antibody was added thereto, followed by room temperature for 15 minutes. cultured in Then, 10 μl of a detection mix containing a cell lysis buffer was added to lyse the cells, followed by reaction at room temperature for 90 minutes. The cell lysate after the reaction was completed was applied to the LANCE cAMP kit (PerkinElmer, USA) to calculate the EC 50 value through the accumulated cAMP, and then compared with each other. Relative titers compared to human GIP are shown in Tables 2 and 3 below.
삼중활성체의 상대적 역가 비율Relative potency ratio of triple active substances
천연형 펩타이드 대비 in vitro 활성 (%) In vitro activity compared to native peptide (%)
서열번호SEQ ID NO: vs GLP-1vs GLP-1 vs Glucagonvs Glucagon vs GIPvs GIP
1One 3.23.2 <0.1<0.1 <0.1<0.1
22 5.95.9 <0.1<0.1 <0.1<0.1
33 1.81.8 <0.1<0.1 <0.1<0.1
44 8.58.5 <0.1<0.1 <0.1<0.1
55 42.142.1 <0.1<0.1 <0.1<0.1
66 17.017.0 <0.1<0.1 <0.1<0.1
77 13.713.7 <0.1<0.1 <0.1<0.1
88 14.214.2 0.100.10 <0.1<0.1
99 32.132.1 0.130.13 <0.1<0.1
1010 46.046.0 <0.1<0.1 <0.1<0.1
1111 1.41.4 <0.1<0.1 <0.1<0.1
1212 0.40.4 <0.1<0.1 <0.1<0.1
1313 < 0.1< 0.1 < 0.1< 0.1 < 0.1< 0.1
1414 28.028.0 < 0.1< 0.1 < 0.1< 0.1
1515 79.279.2 <0.1<0.1 <0.1<0.1
1616 2.12.1 < 0.1< 0.1 < 0.1< 0.1
1717 0.20.2 < 0.1< 0.1 < 0.1< 0.1
1818 <0.1<0.1 <0.1<0.1 <0.1<0.1
1919 <0.1<0.1 <0.1<0.1 <0.1<0.1
2020 <0.1<0.1 <0.1<0.1 <0.1<0.1
2121 17.817.8 267267 22.722.7
2222 20.120.1 140140 59.759.7
2323 4.014.01 9.39.3 <0.1<0.1
2424 41.241.2 9.39.3 < 0.1< 0.1
2525 82.682.6 0.10.1 <0.1<0.1
2626 64.564.5 0.20.2 <0.1<0.1
2727 83.183.1 0.80.8 0.90.9
2828 17.217.2 1.61.6 <0.1<0.1
2929 38.538.5 6.06.0 <0.1<0.1
3030 142142 0.70.7 0.80.8
3131 135135 2.22.2 2.42.4
3232 151151 1.71.7 8.88.8
3333 24.524.5 <0.1<0.1 10.410.4
3434 19.119.1 0.920.92 0.60.6
3535 7.57.5 <0.1<0.1 1.31.3
3636 37.437.4 0.390.39 0.20.2
3737 236236 6.216.21 2.22.2
3838 2.32.3 -- --
3939 13.913.9 0.530.53 <0.1<0.1
4040 75.275.2 <0.1<0.1 <0.1<0.1
4141 34.334.3 <0.1<0.1 <0.1<0.1
4242 33.933.9 205.8205.8 7.87.8
4343 12.612.6 88.488.4 3.703.70
4444 1.31.3 <0.1<0.1 <0.1<0.1
4545 6.66.6 < 0.1< 0.1 < 0.1< 0.1
4646 1.41.4 < 0.1< 0.1 < 0.1< 0.1
4747 2.42.4 < 0.1< 0.1 < 0.1< 0.1
4848 1.51.5 < 0.1< 0.1 < 0.1< 0.1
4949 29.829.8 <0.1<0.1 3.33.3
5050 67.467.4 50.550.5 2.72.7
5151 14.414.4 2.02.0 0.10.1
5252 44.144.1 7.57.5 0.30.3
5353 161161 8.48.4 1.31.3
5454 30.630.6 1.41.4 0.10.1
5555 27.127.1 0.70.7 2.42.4
5656 57.957.9 4.94.9 0.80.8
5757 11.711.7 <0.1<0.1 0.30.3
5858 39.139.1 2.62.6 0.20.2
5959 40.340.3 <0.1<0.1 4.04.0
6060 106.2106.2 <0.1<0.1 8.28.2
6161 59.859.8 <0.1<0.1 2.82.8
6262 5.25.2 <0.1<0.1 <0.1<0.1
6363 15.315.3 <0.1<0.1 <0.1<0.1
6464 64.664.6 60.160.1 92.992.9
6565 95.495.4 25.225.2 11.611.6
6666 15.815.8 172172 17.217.2
6767 28.528.5 46.246.2 39.839.8
6868 27.927.9 8.88.8 107107
6969 24.324.3 9.69.6 62.862.8
7070 15.115.1 71.371.3 64.464.4
7171 90.190.1 12.712.7 94.794.7
7272 11.511.5 1.01.0 1.61.6
7373 22.622.6 5.45.4 3.03.0
7474 12.912.9 0.90.9 1.01.0
7575 35.135.1 8.58.5 18.018.0
7676 10.310.3 47.647.6 11.711.7
7777 38.738.7 12.212.2 35.535.5
7878 51.051.0 14.014.0 0.120.12
7979 41.541.5 4.94.9 1.41.4
8080 8.18.1 0.00.0 0.10.1
8181 7.87.8 0.30.3 <0.1<0.1
8282 9.59.5 1.11.1 <0.1<0.1
8383 47.347.3 1.31.3 0.40.4
8484 4.24.2 <0.1<0.1 <0.1<0.1
8585 4.34.3 <0.1<0.1 0.30.3
8686 28.428.4 0.40.4 0.20.2
8787 0.90.9 <0.1<0.1 <0.1<0.1
8888 9.69.6 0.30.3 <0.1<0.1
8989 7.17.1 0.70.7 <0.1<0.1
9090 7.47.4 <0.1<0.1 <0.1<0.1
9191 31.931.9 16.816.8 0.30.3
9292 0.80.8 <0.1<0.1 0.40.4
9393 5.75.7 0.30.3 0.70.7
9494 0.50.5 <0.1<0.1 <0.1<0.1
9595 2.12.1 0.40.4 <0.1<0.1
9696 34.434.4 194.8194.8 5.25.2
9797 10.510.5 62.862.8 2.62.6
9898 28.128.1 8.28.2 47.147.1
9999 20.920.9 14.914.9 57.757.7
100100 42.242.2 12.712.7 118.5118.5
101101 23.223.2 13.913.9 40.140.1
102102 23.323.3 29.529.5 58.058.0
삼중활성체 지속형 결합체의 상대적 역가 비율Relative potency ratio of triple active long-acting conjugates
지속형 결합체long-acting compound 천연형 펩타이드 대비 in vitro 활성 (%) In vitro activity compared to native peptide (%)
vs GLP-1vs GLP-1 vs Glucagonvs Glucagon vs GIPvs GIP
2121 0.10.1 1.61.6 0.20.2
2222 0.10.1 0.90.9 0.50.5
4242 3.13.1 23.123.1 1.21.2
4343 2.12.1 13.513.5 0.60.6
5050 15.415.4 6.96.9 0.70.7
7777 6.76.7 1.71.7 6.66.6
9696 0.30.3 4.04.0 0.30.3
상기에서 제조한 삼중활성체 또는 이의 지속형 결합체는 GLP-1 수용체, GIP 수용체 및 글루카곤 수용체를 모두 활성화시킬 수 있는 삼중 활성체로 기능을 가지는 바, 목적하는 질환의 치료적 물질로 이용될 수 있다.The triple activator prepared above or a long-acting conjugate thereof has a function as a triple activator capable of activating all of the GLP-1 receptor, the GIP receptor and the glucagon receptor, and thus can be used as a therapeutic substance for a desired disease.
실시예 4: 간 질환 치료 효과 확인Example 4: Confirmation of the treatment effect of liver disease
상기 실시예에서 제조된 삼중 활성체의 지속형 결합체와 FXR (farnesoid X receptor) 작용제, 또는 삼중 활성체의 지속형 결합체와 FXR (farnesoid X receptor) 작용제 및 ACC (acetyla-CoA carboxylase) 억제제 복합투여에 따른 간 질환의 대표적인 예인 NASH 및 섬유증 (fibrosis) 개선효능을 확인코자, NASH 및 fibrosis 모델로 알려진 콜린 결핍, 고지방 및 고콜레스테롤 식이로 유도된 CD-HFD (Choline deficient high fat diet) 마우스 모델을 사용하였다. CD-HFD는 지방간염 및 fibrosis를 유발한다고 알려져 있다. 간략히, C57BL/6 마우스에 8주간 CD-HFD를 진행하여 모델을 유도하였다. The long-acting complex of the triple activator and FXR (farnesoid X receptor) agonist, or the long-acting complex of the triple activator, FXR (farnesoid X receptor) agonist and ACC (acetyla-CoA carboxylase) inhibitor for combined administration of the triple activator prepared in the above Example In order to confirm the efficacy of improving NASH and fibrosis, which are representative examples of liver disease, a choline deficient, high fat and high cholesterol diet, known as NASH and fibrosis model, induced CD-HFD (Choline deficient high fat diet) mouse model was used. . CD-HFD is known to cause steatohepatitis and fibrosis. Briefly, the model was induced by performing CD-HFD in C57BL/6 mice for 8 weeks.
삼중 활성체의 지속형 결합체의 대표예로 서열번호 42의 삼중 활성체의 지속형 결합체, FXR 작용제의 대표예로 킬로펙서 (cilofexor), ACC 억제제의 대표예로 피르소코스타트 (firsocostat)를 선택하여 실험을 수행하였다. 킬로펙서 및 피르소코스타트는 MCE (MedChemExpress)에서 구입하였다.As a representative example of the long-acting complex of the triple activator, a long-acting complex of the triple activator of SEQ ID NO: 42, cilofexor as a representative example of an FXR agonist, and firsocostat as a representative example of an ACC inhibitor Experiments were performed. Kilopexor and pirsocostat were purchased from MCE (MedChemExpress).
4-1: NASH 및 fibrosis 마우스에서 NASH 개선효과 확인4-1: NASH improvement effect confirmed in NASH and fibrosis mice
상기 유도된 동물모델은 부형제 대조군, 서열번호 42의 삼중 활성체의 지속형 결합체 (2.6 nmo/kg, Q2D, 피하) 단독투여군, FXR 작용제 단독 투여군 (4.3 mmol/kg, QD, 경구), ACC 억제제 단독 투여군(51.1 mmol/kg, QD, 경구), 서열번호 42의 삼중 활성체의 지속형 결합체(2.6 nmo/kg, Q2D, 피하)와 FXR 작용제 (4.3 mmol/kg, QD, 경구) 복합투여군, 서열번호 42의 삼중 활성체의 지속형 결합체 (2.6 nmo/kg, Q2D, 피하)와 ACC 억제제(51.1 mmol/kg, QD, 경구) 및 FXR 작용제 (4.3 mmol/kg, QD, 경구) 삼중 복합투여군으로 나누었고, 6주 동안 반복투여를 진행하였다. 6주 반복투여 후 부검을 통해 취한 각 마우스의 간조직에 H&E 염색을 진행한 뒤, NASH 효능평가법으로 잘알려진 NAS (NAFLD activity score)를 구하였다.The induced animal model was an excipient control group, a long-acting conjugate of the triple activator of SEQ ID NO: 42 (2.6 nmo/kg, Q2D, subcutaneous) single administration group, an FXR agonist single administration group (4.3 mmol/kg, QD, oral), ACC inhibitor Single administration group (51.1 mmol/kg, QD, oral), long-acting conjugate of the triple active agent of SEQ ID NO: 42 (2.6 nmo/kg, Q2D, subcutaneous) and FXR agonist (4.3 mmol/kg, QD, oral) combined administration group, Triple active agent of SEQ ID NO: 42 (2.6 nmo/kg, Q2D, subcutaneous) and ACC inhibitor (51.1 mmol/kg, QD, oral) and FXR agonist (4.3 mmol/kg, QD, oral) triple combination administration group , and repeated administration was carried out for 6 weeks. After 6 weeks of repeated administration, H&E staining was performed on the liver tissue of each mouse taken through autopsy, and NAS (NAFLD activity score), which is well-known as a NASH efficacy evaluation method, was obtained.
여기서, QD는 1일 1회, Q2D는 2일에 1회 투여를 의미한다.Here, QD means once a day, and Q2D means once every two days.
도 1에서와 같이 삼중 활성체의 지속형 결합체를 6주간 반복 투여한 결과, CD-HFD 부형제 대조군 대비, NAS가 ACC 억제제 및 FXR 작용제 단독투여군 보다 유의적으로 감소한 것을 확인하였다. 뿐만 아니라, 삼중 활성체의 지속형 결합체를 FXR 작용제와 복합투여 또는 ACC 억제제 및 FXR 작용제와 삼중 복합투여 시, NAS가 보다 효과적으로 개선 된 것을 관찰할 수 있었다.As shown in FIG. 1 , as a result of repeated administration of the triple activator long-acting conjugate for 6 weeks, it was confirmed that, compared to the CD-HFD excipient control group, NAS was significantly reduced compared to the ACC inhibitor and FXR agonist alone group. In addition, it was observed that NAS was more effectively improved when the long-acting conjugate of the triple activator was administered in combination with an FXR agonist or when an ACC inhibitor and FXR agonist were administered in combination.
이를 통해 삼중 활성체 또는 이의 지속형 결합체의 NASH 치료효능이 FXR 작용제와의 복합 투여, 또는 FXR 작용제와 및 ACC 억제제와의 삼중 복합투여로 보다 개선될 수 있음을 확인할 수 있었다.Through this, it was confirmed that the NASH therapeutic efficacy of the triple activator or its long-acting conjugate could be further improved by combined administration with an FXR agonist or triple combination administration with an FXR agonist and an ACC inhibitor.
4-2:4-2: NASH 및 fibrosis 마우스에서 fibrosis 지표 개선효과 확인Confirmation of fibrosis index improvement effect in NASH and fibrosis mice
상기에서 확인한 삼중 활성체의 지속형 결합체 및 FXR 작용제 복합투여, 그리고 삼중 활성체의 지속형 결합체, ACC 억제제 및 FXR 작용제 삼중 복합투여의 fibrosis 치료효능을 확인코자, CD-HFD 마우스 모델의 간조직에서 침윤식 fibrosis 지표로 알려진 hydroxyproline 함량을 측정하였다.In order to confirm the fibrosis treatment efficacy of the long-acting conjugate and FXR agonist combined administration of the triple activator identified above, and the long-acting conjugate of the triple activator, the ACC inhibitor and the triple combination administration of the FXR agonist, in liver tissue of the CD-HFD mouse model The content of hydroxyproline, known as an indicator of infiltrating fibrosis, was measured.
구체적으로, 상기에서 확인한 삼중 활성체의 지속형 결합체의 대표 예인 서열번호 42의 삼중 활성체의 지속형 결합체 단독 투여군, FXR 작용제 단독 투여군, ACC 억제제 단독 투여군, 서열번호 42의 삼중 활성체의 지속형 결합체와 FXR 작용제 복합투여군, 그리고 서열번호 42의 삼중 활성체의 지속형 결합체와 ACC 억제제 및 FXR 작용제 삼중 복합투여군의 CD-HFD 마우스 모델의 간조직에서 hydroxyproline 함량을 측정하였다.Specifically, the long-acting complex of the triple activator of SEQ ID NO: 42, which is a representative example of the long-acting conjugate of the triple activator identified above, the group administered alone, the FXR agonist alone group, the ACC inhibitor alone group, the long-acting type of the triple activator of SEQ ID NO: 42 The hydroxyproline content was measured in the liver tissue of the CD-HFD mouse model of the group administered with the conjugate and the FXR agonist, and the long-acting conjugate of the triple activator of SEQ ID NO: 42, and the triple combination administration of the ACC inhibitor and FXR agonist.
그 결과, 삼중 활성체의 지속형 결합체 투여는 ACC 억제제 및 FXR 작용제 단독투여군 보다 hydroxyproline 함량을 더 감소 시켰다. 또한, 삼중 활성체의 지속형 결합체 투여에 의해 유의적으로 감소한 hydroxyproline 함량이 FXR 작용제와의 복합투여, 그리고 ACC 억제제 및 FXR 작용제 삼중 복합투여시 hydroxyproline 함량이 보다 효과적으로 감소된 것을 관찰할 수 있었다 (도 2).As a result, administration of a long-acting conjugate of the triple activator reduced the hydroxyproline content more than the ACC inhibitor and FXR agonist alone group. In addition, it was observed that the hydroxyproline content significantly decreased by the administration of the long-acting conjugate of the triple activator was more effectively reduced during the combined administration with the FXR agonist and the triple combination administration of the ACC inhibitor and the FXR agonist (Fig. 2).
이를 통해 본 발명의 삼중 활성체 또는 이의 지속형 결합체의 fibrosis 치료효능이 FXR 작용제와의 복합 투여, 또는 FXR 작용제와 및 ACC 억제제와의 삼중 복합투여로 보다 효과적으로 개선될 수 있음을 확인할 수 있었다.Through this, it was confirmed that the fibrosis therapeutic efficacy of the triple activator of the present invention or its long-acting conjugate could be more effectively improved by combined administration with an FXR agonist or triple combination administration with an FXR agonist and an ACC inhibitor.
상기와 같은 결과들은 본 발명의 삼중 활성체 또는 이의 지속형 결합체가 FXR 작용제와의 복합 투여, 또는 FXR 작용제와 및 ACC 억제제와의 삼중 복합투여에 의해 다양한 비알코올성 지방간 질환의 치료 효과를 보다 개선할 수 있음을 시사하는 것으로, 새로운 복합 투여 제제, 또는 삼중 복합 투여 제제로서 이용될 수 있음을 시사하는 것이다.The above results show that the triple active agent or long-acting conjugate of the present invention can further improve the therapeutic effect of various nonalcoholic fatty liver diseases by combined administration with an FXR agonist or triple combination administration with an FXR agonist and an ACC inhibitor. This suggests that it can be used as a new combination dosage formulation, or a triple combination dosage formulation.
이상의 설명으로부터, 본 발명이 속하는 기술분야의 당업자는 본 발명이 그 기술적 사상이나 필수적 특징을 변경하지 않고서 다른 구체적인 형태로 실시될 수 있다는 것을 이해할 수 있을 것이다. 이와 관련하여, 이상에서 기술한 실시 예들은 모든 면에서 예시적인 것이며 한정적인 것이 아닌 것으로서 이해해야만 한다. 본 발명의 범위는 상기 상세한 설명보다는 후술하는 특허 청구범위의 의미 및 범위 그리고 그 등가 개념으로부터 도출되는 모든 변경 또는 변형된 형태가 본 발명의 범위에 포함되는 것으로 해석되어야 한다.From the above description, those skilled in the art to which the present invention pertains will understand that the present invention may be embodied in other specific forms without changing the technical spirit or essential characteristics thereof. In this regard, it should be understood that the embodiments described above are illustrative in all respects and not restrictive. The scope of the present invention should be construed as being included in the scope of the present invention, rather than the above detailed description, all changes or modifications derived from the meaning and scope of the following claims and their equivalents.

Claims (19)

  1. 약학적 유효량의 3중 작용성 지속형 결합체(triple agonistic long-acting conjugate) 또는 3중 작용제와 약학적으로 허용되는 부형제를 함유하는 간 질환의 치료 또는 예방을 위한 약학적 조성물로서,As a pharmaceutical composition for the treatment or prevention of liver disease containing a pharmaceutically effective amount of a triple agonistic long-acting conjugate or a triple agonist and a pharmaceutically acceptable excipient,
    상기 약학적 조성물은 파네소이드 X 수용체 작용제(farnesoid X receptor agonist)와 병용되는 것을 특징으로 하고,The pharmaceutical composition is characterized in that it is used in combination with a farnesoid X receptor agonist,
    상기 3중 작용성 지속형 결합체는 하기 화학식 1로 표시되는 물질인 약학적 조성물: The triple-acting long-acting conjugate is a pharmaceutical composition of a substance represented by the following formula (1):
    [화학식 1][Formula 1]
    Z - Lx - FcZ - Lx - Fc
    이 때 Lx는 에틸렌글리콜 반복 단위를 함유하는 링커이며, x는 0 또는 자연수이고,In this case, Lx is a linker containing an ethylene glycol repeating unit, and x is 0 or a natural number,
    Fc는 면역글로불린 Fc 영역이고, Fc is an immunoglobulin Fc region,
    -는 Lx와 Z 사이, Fc와 Lx 사이의 각각에 공유결합 연결을 나타내며,- represents a covalent linkage between Lx and Z, between Fc and Lx, respectively,
    Z 또는 3중 작용제는 하기 일반식 1로 표시되는 아미노산 서열을 포함하는 펩타이드이고, Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 1,
    Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1(일반식 1, 서열번호 103)Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp23 Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (Formula 1, SEQ ID NO: 103)
    상기 일반식 1에서,In the above general formula 1,
    Xaa1은 히스티딘(His, H), 4-이미다조아세틸(CA) 또는 티로신(Tyr, Y)이고, Xaa1 is histidine (His, H), 4-imidazoacetyl (CA) or tyrosine (Tyr, Y),
    Xaa2는 글리신(Gly, G), 알파-메틸-글루탐산 또는 2-아미노이소부티르산(2-aminoisobutyric acid)이며,Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid or 2-aminoisobutyric acid,
    Xaa3은 글루탐산(Glu, E) 또는 글루타민(Gln, Q)이고, Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q),
    Xaa7은 트레오닌(Thr, T) 또는 이소류신(Ile, I)이며,Xaa7 is threonine (Thr, T) or isoleucine (Ile, I),
    Xaa10은 류신(Leu, L), 티로신(Tyr, Y), 리신(Lys, K), 시스테인(Cys, C) 또는 발린(Val, V)이고, Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C) or valine (Val, V);
    Xaa12는 리신(Lys, K), 세린(Ser, S) 또는 이소류신(Ile, I)이며, Xaa12 is lysine (Lys, K), serine (Ser, S) or isoleucine (Ile, I),
    Xaa13은 글루타민(Gln, Q), 티로신(Tyr, Y), 알라닌(Ala, A) 또는 시스테인(Cys, C)이고, Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A) or cysteine (Cys, C);
    Xaa14는 류신(Leu, L), 메티오닌(Met, M) 또는 티로신(Tyr, Y)이며,Xaa14 is leucine (Leu, L), methionine (Met, M) or tyrosine (Tyr, Y);
    Xaa15는 시스테인(Cys, C), 아스파르트산(Asp, D), 글루탐산(Glu, E) 또는 류신(Leu, L)이며,Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E) or leucine (Leu, L);
    Xaa16은 글리신(Gly, G), 글루탐산(Glu, E) 또는 세린(Ser, S)이고, Xaa16 is glycine (Gly, G), glutamic acid (Glu, E) or serine (Ser, S);
    Xaa17은 글루타민(Gln, Q), 아르기닌(Arg, R), 이소류신(Ile, I), 글루탐산(Glu, E), 시스테인(Cys, C) 또는 리신(Lys, K)이며,Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C) or lysine (Lys, K);
    Xaa18은 알라닌(Ala, A), 글루타민(Gln, Q), 아르기닌(Arg, R) 또는 히스티딘(His, H)이고,Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R) or histidine (His, H);
    Xaa19는 알라닌(Ala, A), 글루타민(Gln, Q), 시스테인(Cys, C) 또는 발린(Val, V)이며,Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C) or valine (Val, V);
    Xaa20은 리신(Lys, K), 글루타민(Gln, Q) 또는 아르기닌(Arg, R)이고,Xaa20 is lysine (Lys, K), glutamine (Gln, Q) or arginine (Arg, R);
    Xaa21은 글루탐산(Glu, E), 글루타민(Gln, Q), 류신(Leu, L), 시스테인(Cys, C) 또는 아스파르트산(Asp, D)이며,Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C) or aspartic acid (Asp, D);
    Xaa23은 이소류신(Ile, I) 또는 발린(Val, V)이고,Xaa23 is isoleucine (Ile, I) or valine (Val, V),
    Xaa24는 알라닌(Ala, A), 글루타민(Gln, Q), 시스테인(Cys, C), 아스파라긴(Asn, N), 아스파르트산(Asp, D) 또는 글루탐산(Glu, E)이며,Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D) or glutamic acid (Glu, E),
    Xaa27은 발린(Val, V), 류신(Leu, L) 또는 리신(Lys, K) 이고,Xaa27 is valine (Val, V), leucine (Leu, L) or lysine (Lys, K);
    Xaa28은 시스테인(Cys, C), 리신(Lys, K), 알라닌(Ala, A), 아스파라긴(Asn, N) 또는 아스파르트산(Asp, D)이며,Xaa28 is cysteine (Cys, C), lysine (Lys, K), alanine (Ala, A), asparagine (Asn, N) or aspartic acid (Asp, D);
    Xaa29는 시스테인(Cys, C), 글리신(Gly, G), 글루타민(Gln, Q), 트레오닌(Thr, T), 글루탐산(Glu, E) 또는 히스티딘(His, H)이고,Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q), threonine (Thr, T), glutamic acid (Glu, E) or histidine (His, H);
    Xaa30은 시스테인(Cys, C), 글리신(Gly, G), 리신(Lys, K) 또는 히스티딘(His, H)이거나, 부존재하며,Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K) or histidine (His, H), or is absent;
    R1은 시스테인(Cys, C), GKKNDWKHNIT(서열번호 106), m-SSGAPPPS-n(서열번호 107) 또는 m-SSGQPPPS-n(서열번호 108)이거나, 부존재하며, R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107) or m-SSGQPPPS-n (SEQ ID NO: 108), or is absent;
    m은 -Cys-, -Pro- 또는 -Gly-Pro-이고, m is -Cys-, -Pro- or -Gly-Pro-;
    n은 부존재하거나 -Cys-, -Gly-, -Ser- 또는 -His-Gly-이다.n is absent or is -Cys-, -Gly-, -Ser- or -His-Gly-.
  2. 제1항에 있어서, The method of claim 1,
    Xaa2는 글리신, 알파-메틸-글루탐산 또는 2-아미노이소부티르산이고,Xaa2 is glycine, alpha-methyl-glutamic acid or 2-aminoisobutyric acid,
    Xaa7은 트레오닌이며,Xaa7 is threonine,
    Xaa10은 티로신, 시스테인 또는 발린이고, Xaa10 is tyrosine, cysteine or valine,
    Xaa12는 리신 또는 이소류신이며, Xaa12 is lysine or isoleucine,
    Xaa13은 티로신, 알라닌, 글루타민 또는 시스테인이고, Xaa13 is tyrosine, alanine, glutamine or cysteine,
    Xaa14는 류신, 티로신 또는 메티오닌이며;Xaa14 is leucine, tyrosine or methionine;
    Xaa15는 시스테인, 류신, 글루탐산 또는 아스파르트산이고,Xaa15 is cysteine, leucine, glutamic acid or aspartic acid,
    Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 글루탐산 또는 리신이며,Xaa17 is glutamine, arginine, isoleucine, cysteine, glutamic acid or lysine,
    Xaa18은 알라닌, 글루타민, 아르기닌 또는 히스티딘이고,Xaa18 is alanine, glutamine, arginine or histidine;
    Xaa19는 알라닌, 글루타민, 발린 또는 시스테인이며,Xaa19 is alanine, glutamine, valine or cysteine,
    Xaa20은 리신, 아르기닌 또는 글루타민이고,Xaa20 is lysine, arginine or glutamine,
    Xaa21은 글루탐산, 글루타민, 류신, 시스테인 또는 아스파르트산이며,Xaa21 is glutamic acid, glutamine, leucine, cysteine or aspartic acid,
    Xaa23은 이소류신 또는 발린이고,Xaa23 is isoleucine or valine,
    Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 글루탐산 또는 아스파르트산이며,Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid or aspartic acid;
    Xaa27은 류신 또는 리신인 약학적 조성물.Xaa27 is leucine or lysine.
  3. 제1항에 있어서, Z 또는 3중 작용제는 하기 일반식 2로 표시되는 아미노산 서열을 포함하는 펩타이드인 약학적 조성물:The pharmaceutical composition according to claim 1, wherein the Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 2:
    Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31- Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40(일반식 2, 서열번호 104)Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (Formula 2, SEQ ID NO: 104)
    상기 일반식 2에서,In the above general formula 2,
    Xaa1은 4-이미다조아세틸, 히스티딘 또는 티로신이고,Xaa1 is 4-imidazoacetyl, histidine or tyrosine;
    Xaa2는 글리신, 알파-메틸-글루탐산 또는 2-아미노이소부티르산이며,Xaa2 is glycine, alpha-methyl-glutamic acid or 2-aminoisobutyric acid;
    Xaa10은 티로신 또는 시스테인이고,Xaa10 is tyrosine or cysteine,
    Xaa13은 알라닌, 글루타민, 티로신 또는 시스테인이며,Xaa13 is alanine, glutamine, tyrosine or cysteine,
    Xaa14는 류신, 메티오닌 또는 티로신이고,Xaa14 is leucine, methionine or tyrosine,
    Xaa15는 아스파르트산, 글루탐산 또는 류신이며,Xaa15 is aspartic acid, glutamic acid or leucine,
    Xaa16은 글리신, 글루탐산 또는 세린이고,Xaa16 is glycine, glutamic acid or serine,
    Xaa17은 글루타민, 아르기닌, 이소류신, 글루탐산, 시스테인 또는 리신이며,Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine or lysine,
    Xaa18은 알라닌, 글루타민, 아르기닌 또는 히스티딘이고,Xaa18 is alanine, glutamine, arginine or histidine;
    Xaa19는 알라닌, 글루타민, 시스테인 또는 발린이며,Xaa19 is alanine, glutamine, cysteine or valine,
    Xaa20은 리신, 글루타민 또는 아르기닌이고,Xaa20 is lysine, glutamine or arginine,
    Xaa21은 시스테인, 글루탐산, 글루타민, 류신 또는 아스파르트산이며,Xaa21 is cysteine, glutamic acid, glutamine, leucine or aspartic acid,
    Xaa23은 이소류신 또는 발린이고,Xaa23 is isoleucine or valine,
    Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴 또는 글루탐산이며,Xaa24 is cysteine, alanine, glutamine, asparagine or glutamic acid,
    Xaa28은 리신, 시스테인, 아스파라긴 또는 아스파르트산이고,Xaa28 is lysine, cysteine, asparagine or aspartic acid;
    Xaa29는 글리신, 글루타민, 시스테인 또는 히스티딘이며,Xaa29 is glycine, glutamine, cysteine or histidine;
    Xaa30은 시스테인, 글리신, 리신 또는 히스티딘이고,Xaa30 is cysteine, glycine, lysine or histidine;
    Xaa31은 프롤린 또는 시스테인이며;Xaa31 is proline or cysteine;
    Xaa40은 시스테인이거나, 부존재한다.Xaa40 is cysteine or absent.
  4. 제1항에 있어서, Z 또는 3중 작용제는 하기 일반식 3으로 표시되는 아미노산 서열을 포함하는 펩타이드인 약학적 조성물:The pharmaceutical composition according to claim 1, wherein the Z or triple agent is a peptide comprising an amino acid sequence represented by the following general formula 3:
    Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40(일반식 3, 서열번호 105),Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp- Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (general formula 3, SEQ ID NO: 105);
    상기 일반식 3에서,In the above general formula 3,
    Xaa1은 히스티딘 또는 티로신이고,Xaa1 is histidine or tyrosine,
    Xaa2는 알파-메틸-글루탐산 또는 2-아미노이소부티르산이며,Xaa2 is alpha-methyl-glutamic acid or 2-aminoisobutyric acid,
    Xaa13은 알라닌, 티로신 또는 시스테인이고,Xaa13 is alanine, tyrosine or cysteine,
    Xaa17은 아르기닌, 시스테인 또는 리신이며,Xaa17 is arginine, cysteine or lysine,
    Xaa18은 알라닌 또는 아르기닌이고,Xaa18 is alanine or arginine,
    Xaa19는 알라닌 또는 시스테인이며,Xaa19 is alanine or cysteine,
    Xaa21은 글루탐산 또는 아스파르트산이고,Xaa21 is glutamic acid or aspartic acid,
    Xaa24는 글루타민 또는 아스파라긴이며,Xaa24 is glutamine or asparagine,
    Xaa28은 시스테인 또는 아스파르트산이고,Xaa28 is cysteine or aspartic acid,
    Xaa29는 시스테인, 히스티딘 또는 글루타민이며,Xaa29 is cysteine, histidine or glutamine;
    Xaa30은 시스테인 또는 히스티딘이고,Xaa30 is cysteine or histidine,
    Xaa31은 프롤린 또는 시스테인이며,Xaa31 is proline or cysteine,
    Xaa40은 시스테인 또는 부존재함.Xaa40 is cysteine or absent.
  5. 제1항에 있어서, 상기 Xaa16이 글루탐산이고, Xaa20이 리신이며, 상기 글루탐산과 상기 리신이 락탐 고리를 형성한 약학적 조성물.The pharmaceutical composition according to claim 1, wherein the Xaa16 is glutamic acid, Xaa20 is lysine, and the glutamic acid and the lysine form a lactam ring.
  6. 제1항에 있어서, 상기 Z 또는 3중 작용제는 그 C-말단이 아미드화된 약학적 조성물.The pharmaceutical composition according to claim 1, wherein the Z or triple agent is amidated at its C-terminus.
  7. 제1항에 있어서, 상기 Lx 내의 에틸렌글리콜 반복 단위 부분의 화학식량은 1 ~ 100 kDa 범위에 있는 약학적 조성물.The pharmaceutical composition according to claim 1, wherein the formula weight of the ethylene glycol repeating unit moiety in Lx is in the range of 1 to 100 kDa.
  8. 제1항에 있어서, Z 또는 3중 작용제는 서열번호: 1 내지 102로 이루어진 군으로부터 선택하는 어느 하나의 아미노산 서열을 포함하는 펩타이드인 약학적 조성물.The pharmaceutical composition according to claim 1, wherein the Z or triple agent is a peptide comprising any one amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102.
  9. 제8항에 있어서, Z 또는 3중 작용제는 서열번호 21, 22, 42, 43, 50, 64, 66, 67, 70, 71, 76, 77, 96, 97과 100으로 이루어진 군으로부터 선택하는 아미노산 서열을 포함하는 펩타이드인 약학적 조성물.9. The method of claim 8, wherein the Z or triple agent is an amino acid selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 64, 66, 67, 70, 71, 76, 77, 96, 97 and 100. A pharmaceutical composition which is a peptide comprising a sequence.
  10. 제9항에 있어서, 상기 펩타이드는 서열번호 21, 22, 42, 43, 50, 66, 67, 77, 96, 97과 100으로 이루어진 군으로부터 선택하는 아미노산 서열을 포함하는 펩타이드인 약학적 조성물.The pharmaceutical composition according to claim 9, wherein the peptide is a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 66, 67, 77, 96, 97 and 100.
  11. 제10항에 있어서, 상기 펩타이드는 서열번호 21, 22, 42, 43, 50, 77과 96으로 이루어진 군으로부터 선택하는 아미노산 서열을 포함하는 펩타이드인 약학적 조성물.The pharmaceutical composition according to claim 10, wherein the peptide is a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 77 and 96.
  12. 제1항에 있어서, 상기 파네소이드 X 수용체 작용제는 카페스톨(Cafestol), 케노데옥시콜린산(Chenodeoxycholic acid), 오베티콜린산(Obeticholic acid), 펙사라민(Fexaramine), GW 4064, PX104, 6E-CDCA(6-ethyl-chedeoxycholic acid), AKN-083, 트로피펙서(Tropifexor), 킬로펙서(Cilofexor), EDP-305 AGN-242266, AGN-242256, EP-024297, RDX-023, BWL-200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, PX20606, EYP-001, TERN-101, TC-100 및 INT-2228으로 이루어진 군에서 선택하는 약학적 조성물.The method of claim 1, wherein the farnesoid X receptor agonist is cafestol, chenodeoxycholic acid, obeticholic acid, fexaramine, GW 4064, PX104 , 6E-CDCA (6-ethyl-chedeoxycholic acid), AKN-083, Tropifexor, Cilofexor, EDP-305 AGN-242266, AGN-242256, EP-024297, RDX-023, BWL- 200, GNF-5120, GS-9674, LMB-763, Px-102, Px-103, M790, M780, M450, M-480, MET-409, MET-642, PX20606, EYP-001, TERN-101, A pharmaceutical composition selected from the group consisting of TC-100 and INT-2228.
  13. 제1항 내지 제12항 중 어느 한 항에 있어서, 상기 약학적 조성물은 아세틸조효소 A 카르복실라제 억제제(acetyl-Coenzyme A carboxylase inhibitor)와도 병용하는 것을 특징으로 하는 약학적 조성물.The pharmaceutical composition according to any one of claims 1 to 12, wherein the pharmaceutical composition is also used in combination with an acetyl-Coenzyme A carboxylase inhibitor.
  14. 제13항에 있어서, 아세틸조효소 A 카르복실라제 억제제는 CP-640186,(4-피페리디닐)-피페라진 유도체((4-piperidinyl)-piperazine derivatives), 1,4-치환 사이클로헥산 유도체(1,4-disubstituted cyclohexane derivatives), 스피로크로마논 유도체(Spirochromanone derivatives), 스피로락탐 유도체(Spirolactam derivatives), 스피로디아민 유도체(Spirodiamine derivatives), 스피로펜타실아미드 유도체(Spiropentacylamide derivatives), 슈도펩타이드 피롤리딘다이온 유도체(Pseudopeptide pyrrolidinedione derivatives), 매크로사이클릭 폴리케톤 유도체(Macrocyclic polyketone derivatives), 피르소코스타트 (firsocostat)를 포함하는 티오펜 피리미돈 유도체(Thiophene pyrimidone derivatives), 아미노-옥사졸 유도체(Amino-oxazole derivatives), 아조벤지미다졸 유도체(Azobenzimidazole derivatives) 및 PF-05221304로 이루어진 군에서 선택하는 약학적 조성물.14. The method of claim 13, wherein the acetyl coenzyme A carboxylase inhibitor is CP-640186, (4-piperidinyl)-piperazine derivatives ((4-piperidinyl)-piperazine derivatives), 1,4-substituted cyclohexane derivatives (1 ,4-disubstituted cyclohexane derivatives, Spirochromanone derivatives, Spirolactam derivatives, Spirodiamine derivatives, Spiropentacylamide derivatives, Pseudopeptide pyrrolidinedione derivatives (Pseudopeptide pyrrolidinedione derivatives), Macrocyclic polyketone derivatives, Thiophene pyrimidone derivatives including firsocostat, Amino-oxazole derivatives, A pharmaceutical composition selected from the group consisting of Azobenzimidazole derivatives and PF-05221304.
  15. 제1항 내지 제12항 중 어느 한 항에 있어서, 상기 간 질환은 비알코올성 지방간 질환(non-alcoholic fatty liver disease) 또는 담즙정체성 간질환인 것인 약학적 조성물.The pharmaceutical composition according to any one of claims 1 to 12, wherein the liver disease is non-alcoholic fatty liver disease or cholestatic liver disease.
  16. 제15항에 있어서, 상기 비알코올성 지방간 질환은 단순 지방증, 간 염증, 비알코올성 지방간, 비알코올성 지방간염, 간경변, 간 섬유증, 간부전, 및 간암으로 이루어진 군에서 선택하는 적어도 하나의 질환인 약학적 조성물.The pharmaceutical composition according to claim 15, wherein the nonalcoholic fatty liver disease is at least one disease selected from the group consisting of simple steatosis, liver inflammation, nonalcoholic fatty liver, nonalcoholic steatohepatitis, cirrhosis, liver fibrosis, liver failure, and liver cancer. .
  17. 제15항에 있어서, 상기 담즙 정체성 간질환은 원발성 담즙성 경변증 (Primary biliary cirrhosis), 원발성 경화성 담관염(primary sclerosing cholangitis) 및 이들의 조합으로 이루어진 군에서 선택하는 어느 하나인 약학적 조성물.The pharmaceutical composition according to claim 15, wherein the cholestatic liver disease is any one selected from the group consisting of primary biliary cirrhosis, primary sclerosing cholangitis, and combinations thereof.
  18. 제15항 내지 제17항 중 어느 한 항에 있어서, 상기 약학적 조성물은 아세틸조효소 A 카르복실라제 억제제(acetyl-Coenzyme A carboxylase inhibitor)와도 병용하는 것을 특징으로 하는 약학적 조성물.The pharmaceutical composition according to any one of claims 15 to 17, wherein the pharmaceutical composition is also used in combination with an acetyl-Coenzyme A carboxylase inhibitor.
  19. 제15항에 있어서, 상기 비알코올성 지방간 질환은 간 염증, 비알코올성 지방간염과 간 섬유증으로 이루어진 군에서 선택하는 적어도 하나의 질환인 약학적 조성물.The pharmaceutical composition according to claim 15, wherein the nonalcoholic fatty liver disease is at least one disease selected from the group consisting of liver inflammation, nonalcoholic steatohepatitis and liver fibrosis.
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